Egfr nucleic acids and uses thereof

ABSTRACT

Disclosed herein are molecules and pharmaceutical compositions that mediate RNA interference against EGFR. Also described herein include methods for treating a disease or disorder that comprises a molecule or a pharmaceutical composition that mediate RNA interference against EGFR.

CROSS-REFERENCE

This application claims the benefit of U.S. Provisional Application No.62/317,105, filed Apr. 1, 2016, which application is incorporated hereinby reference.

SEQUENCE LISTING

The instant application contains a Sequence Listing which has beensubmitted electronically in ASCII format and is hereby incorporated byreference in its entirety. Said ASCII copy, created on Mar. 24, 2017, isnamed 45532-709_201_SL.txt and is 554,437 bytes in size.

BACKGROUND OF THE DISCLOSURE

Gene suppression by RNA-induced gene silencing provides several levelsof control: transcription inactivation, small interfering RNA(siRNA)-induced mRNA degradation, and siRNA-induced transcriptionalattenuation. In some instances, RNA interference (RNAi) provides longlasting effect over multiple cell divisions. As such, RNAi represents aviable method useful for drug target validation, gene function analysis,pathway analysis, and disease therapeutics.

SUMMARY OF THE DISCLOSURE

Disclosed herein, in certain embodiments, are molecules andpharmaceutical compositions for modulating EGFR function and/orexpression in a cell.

Disclosed herein, in certain embodiments, is a polynucleic acid moleculethat mediates RNA interference against EGFR, wherein the polynucleicacid molecule comprises at least one 2′ modified nucleotide, at leastone modified internucleotide linkage, or at least one inverted abasicmoiety.

In some embodiments, the at least one 2′ modified nucleotide comprises2′-O-methyl, 2′-O-methoxyethyl (2′-O-MOE), 2′-O-aminopropyl, 2′-deoxy,T-deoxy-2′-fluoro, 2′-O-aminopropyl (2′-O-AP), 2′-O-dimethylaminoethyl(2′-O-DMAOE), 2′-O-dimethylaminopropyl (2′-O-DMAP),T-O-dimethylaminoethyloxyethyl (2′-O-DMAEOE), or 2′-O—N-methylacetamido(2′-O-NMA) modified nucleotide. In some embodiments, the at least one 2′modified nucleotide comprises locked nucleic acid (LNA) or ethylenenucleic acid (ENA). In some embodiments, the at least one inverted basicmoiety is at at least one terminus. In some embodiments, the at leastone modified internucleotide linkage comprises a phosphorothioatelinkage or a phosphorodithioate linkage.

In some embodiments, the polynucleic acid molecule is at least fromabout 10 to about 30 nucleotides in length. In some embodiments, thepolynucleic acid molecule is at least one of: from about 15 to about 30,from about 18 to about 25, form about 18 to about 24, from about 19 toabout 23, or from about 20 to about 22 nucleotides in length. In someembodiments, the polynucleic acid molecule is at least about 16, 17, 18,19, 20, 21, 22, 23, 24, or 25 nucleotides in length.

In some embodiments, the polynucleic acid molecule comprises at leastone of: from about 5% to about 100% modification, from about 10% toabout 100% modification, from about 20% to about 100% modification, fromabout 30% to about 100% modification, from about 40% to about 100%modification, from about 50% to about 100% modification, from about 60%to about 100% modification, from about 70% to about 100% modification,from about 80% to about 100% modification, and from about 90% to about100% modification.

In some embodiments, the polynucleic acid molecule comprises at leastone of: from about 10% to about 90% modification, from about 20% toabout 90% modification, from about 30% to about 90% modification, fromabout 40% to about 90% modification, from about 50% to about 90%modification, from about 60% to about 90% modification, from about 70%to about 90% modification, and from about 80% to about 100%modification.

In some embodiments, the polynucleic acid molecule comprises at leastone of: from about 10% to about 80% modification, from about 20% toabout 80% modification, from about 30% to about 80% modification, fromabout 40% to about 80% modification, from about 50% to about 80%modification, from about 60% to about 80% modification, and from about70% to about 80% modification.

In some embodiments, the polynucleic acid molecule comprises at leastone of: from about 10% to about 70% modification, from about 20% toabout 70% modification, from about 30% to about 70% modification, fromabout 40% to about 70% modification, from about 50% to about 70%modification, and from about 60% to about 70% modification.

In some embodiments, the polynucleic acid molecule comprises at leastone of: from about 10% to about 60% modification, from about 20% toabout 60% modification, from about 30% to about 60% modification, fromabout 40% to about 60% modification, and from about 50% to about 60%modification.

In some embodiments, the polynucleic acid molecule comprises at leastone of: from about 10% to about 50% modification, from about 20% toabout 50% modification, from about 30% to about 50% modification, andfrom about 40% to about 50% modification.

In some embodiments, the polynucleic acid molecule comprises at leastone of: from about 10% to about 40% modification, from about 20% toabout 40% modification, and from about 30% to about 40% modification.

In some embodiments, the polynucleic acid molecule comprises at leastone of: from about 10% to about 30% modification, and from about 20% toabout 30% modification.

In some embodiments, the polynucleic acid molecule comprises from about10% to about 20% modification.

In some embodiments, the polynucleic acid molecule comprises from about15% to about 90%, from about 20% to about 80%, from about 30% to about70%, or from about 40% to about 60% modifications.

In some embodiments, the polynucleic acid molecule comprises at leastabout 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99%modification.

In some embodiments, the polynucleic acid molecule comprises at leastabout 3, about 4, about 5, about 6, about 7, about 8, about 9, about 10,about 11, about 12, about 13, about 14, about 15, about 16, about 17,about 18, about 19, about 20, about 21, about 22 or more modifications.

In some embodiments, the polynucleic acid molecule comprises at leastabout 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8,about 9, about 10, about 11, about 12, about 13, about 14, about 15,about 16, about 17, about 18, about 19, about 20, about 21, about 22 ormore modified nucleotides.

In some embodiments, the polynucleic acid molecule comprises a sequencethat hybridizes to a target sequence selected from SEQ ID NOs: 1-376.

In some embodiments, the polynucleic acid molecule comprises a singlestrand.

In some embodiments, the polynucleic acid molecule comprises two or morestrands.

In some embodiments, the polynucleic acid molecule comprises a firstpolynucleotide and a second polynucleotide hybridized to the firstpolynucleotide to form a double-stranded polynucleic acid molecule.

In some embodiments, the second polynucleotide comprises at least onemodification.

In some embodiments, the first polynucleotide and the secondpolynucleotide are RNA molecules. In some embodiments, the firstpolynucleotide and the second polynucleotide are siRNA molecules.

In some embodiments, the first polynucleotide comprises a sequencehaving at least 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%,99%, or 100% sequence identity to SEQ ID NOs: 377-1892. In someembodiments, the first polynucleotide consists of a sequence selectedfrom SEQ ID NOs: 377-1892. In some embodiments, the secondpolynucleotide comprises a sequence having at least 60%, 65%, 70%, 75%,80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to SEQID NOs: 377-1892. In some embodiments, the second polynucleotideconsists of a sequence selected from SEQ ID NOs: 377-1892.

Disclosed herein, in certain embodiments, is a pharmaceuticalcomposition comprising: a) a molecule disclosed above; and b) apharmaceutically acceptable excipient. In some embodiments, thepharmaceutical composition is formulated as a nanoparticle formulation.In some embodiments, the pharmaceutical composition is formulated forparenteral, oral, intranasal, buccal, rectal, or transdermaladministration.

Disclosed herein, in certain embodiments, is a method of treating adisease or disorder in a patient in need thereof, comprisingadministering to the patient a composition comprising a moleculedisclosed above. In some embodiments, the disease or disorder is acancer. In some embodiments, the cancer is a solid tumor. In someembodiments, the cancer is a hematologic malignancy. In someembodiments, the cancer comprises an EGFR-associated cancer. In someembodiments, the cancer comprises bladder cancer, breast cancer,colorectal cancer, endometrial cancer, esophageal cancer, glioblastomamultiforme, head and neck cancer, kidney cancer, lung cancer, ovariancancer, pancreatic cancer, prostate cancer, or thyroid cancer. In someembodiments, the cancer comprises acute myeloid leukemia, CLL, DLBCL, ormultiple myeloma.

Disclosed herein, in certain embodiments, is a method of inhibiting theexpression of a target gene in a primary cell of a patient, comprisingadministering a molecule disclosed above to the primary cell. In someembodiments, the method is an in vivo method. In some embodiments, thepatient is a human.

Disclosed herein, in certain embodiments, is a kit comprising a moleculedisclosed above.

DETAILED DESCRIPTION OF THE DISCLOSURE

Epidermal growth factor receptor (EGFR, ErbB-1, or HER1) is atransmembrane tyrosine kinase receptor and a member of the ErbB familyof receptors, which also include HER2/c-neu (ErbB-2), Her3 (ErbB-3), andHer4 (ErbB-4). In some instances, EGFR mutations drive the downstreamactivation of RAS/RAF/MAPK, PI3K/AKT, and/or JAK/STAT pathways, leadingto mitosis, cell proliferation, and suppression of apoptosis. Inaddition, amplification of wild-type EGFR gene has been implicated inthe development of cancers such as glioblastomas and non-small cell lungcancer (Talasila, et al., “EGFR Wild-type Amplification and ActivationPromote Invasion and Development of Glioblastoma Independent ofAngiogenesis,” Acta Neuropathol. 125(5): 683-698 (2013); Bell et al.,“Epidermal Growth Factor Receptor Mutations and Gene Amplification inNon-Small-Cell Lung Cancer: Molecular Analysis of the IDEAL/INTACTGefitinib Trials,” J. Clinical Oncology 23(31): 8081-8092 (2005)).

Disclosed herein, in certain embodiments, are polynucleic acid moleculesand pharmaceutical compositions that modulate the expression of EGFR. Insome instances, the polynucleic acid molecules and pharmaceuticalcompositions modulate the expression of wild type EGFR gene. In otherinstances, the polynucleic acid molecules and pharmaceuticalcompositions modulate the expression of mutant EGFR.

In some embodiments, the polynucleic acid molecules and pharmaceuticalcompositions are used for the treatment of a disease or disorder (e.g.,cancer or an EGFR-associated disease or disorder). In additionalembodiments, the polynucleic acid molecules and pharmaceuticalcompositions are used for inhibiting the expression of EGFR gene in aprimary cell of a patient in need thereof.

In additional cases, also included herein are kits that comprise one ormore of polynucleic acid molecules and pharmaceutical compositionsdescribed herein.

Polynucleic Acid Molecule

In some embodiments, a polynucleic acid molecule described hereinmodulates the expression of the EGFR gene (GenBank: BC094761.1). In someembodiments, EGFR DNA or RNA is wild type EGFR or EGFR comprising amutation. In some instances, EGFR is wild type EGFR. In some instances,EGFR DNA or RNA comprises a mutation. In some instances, the polynucleicacid molecule hybridizes to a target region of wild type EGFR DNA orRNA. In some instances, the polynucleic acid molecule hybridizes to atarget region of EGFR DNA or RNA comprising a mutation (e.g., asubstitution, a deletion, or an addition).

In some instances, EGFR DNA or RNA comprises one or more mutations. Insome embodiments, EGFR DNA or RNA comprises one or more mutations withinone or more exons. In some instances, the one or more exons compriseexon 18, exon 19, exon 20, exon 21, or exon 22. In some instances, EGFRDNA or RNA comprises one or more mutations in exon 18, exon 19, exon 20,exon 21, exon 22, or a combination thereof.

In some instances, EGFR DNA or RNA comprises one or more mutations atpositions corresponding to amino acid residues 34, 38, 45, 62, 63, 77,78, 108, 114, 120, 140, 148, 149, 160, 177, 178, 189, 191, 198, 220,222, 223, 229, 237, 240, 244, 252, 254, 255, 256, 263, 270, 273, 276,282, 288, 289, 301, 303, 304, 309, 314, 326, 331, 354, 363, 373, 337,380, 384, 393, 427, 428, 437, 441, 447, 465, 475, 515, 526, 527, 531,536, 541, 546, 571, 588, 589, 596, 596, 598, 602, 614, 620, 628, 636,641, 645, 651, 671, 689, 694, 700, 709, 712, 714, 715, 716, 719, 720,721, 731, 733, 739-744, 742, 746-750, 746-752, 746, 747, 747-749,747-751, 747-753, 751, 752, 754, 752-759, 750, 761-762, 761, 763, 765,767-768, 767-769, 768, 769, 769-770, 770-771, 772, 773-774, 773, 774,774-775, 776, 779, 783, 784, 786, 790, 792, 794, 798, 803, 805, 807,810, 826, 827, 831, 832, 833, 835, 837, 838, 839, 842, 843, 847, 850,851, 853, 854, 856, 858, 861, 863, 894, 917, 967, 1006, 1019, 1042,1100, 1129, 1141, 1153, 1164, 1167, or a combination thereof of the EGFRpolypeptide. In some embodiments, EGFR DNA or RNA comprises one or moremutations at positions corresponding to amino acid residues 747, 761,790, 854, 858, or a combination thereof of the EGFR polypeptide. In someembodiments, EGFR DNA or RNA comprises one or more mutations atpositions corresponding to amino acid residues 761, 790, 858, or acombination thereof of the EGFR polypeptide. In some embodiments, EGFRDNA or RNA comprises a mutation at a position corresponding to aminoacid residue 747 of the EGFR polypeptide. In some embodiments, EGFR DNAor RNA comprises a mutation at a position corresponding to amino acidresidue 761 of the EGFR polypeptide. In some embodiments, EGFR DNA orRNA comprises a mutation at a position corresponding to amino acidresidue 790 of the EGFR polypeptide. In some embodiments, EGFR DNA orRNA comprises a mutation at a position corresponding to amino acidresidue 854 of the EGFR polypeptide. In some embodiments, EGFR DNA orRNA comprises a mutation at a position corresponding to amino acidresidue 858 of the EGFR polypeptide.

In some embodiments, EGFR DNA or RNA comprises one or more mutationsselected from T34M, L38V, E45Q, L62R, G63R, G63K, S77F, F78L, R108K,R108G, E114K, A120P, L140V, V148M, R149W, E160K, S177P, M178I, K189T,D191N, S198R, S220P, R222L, R222C, S223Y, S229C, A237Y, C240Y, R244G,R252C, R252P, F254I, R255 (nonsense mutation), D256Y, T263P, Y270C,T273A, Q276 (nonsense), E282K, G288 (frame shift), A289D, A289V, A289T,A289N, A289D, V301 (deletion), D303H, H304Y, R309Q, D314N, C326R, G331R,T354M, T363I, P373Q, R337S, S380 (frame shift), T384S, D393Y, R427L,G428S, S437Y, V441I, S447Y, G465R, I475V, C515S, C526S, R527L, R531(nonsense), V536M, L541I, P546Q, C571S, G588S, P589L, P596L, P596S,P596R, P596L, G598V, G598A, E602G, G614D, C620Y, C620W, C628Y, C628F,C636Y, T638M, P641H, S645C, V651M, R671C, V689M, P694S, N700D, E709A,E709K, E709Q, E709K, F712L, K714N, I715S, K716R, G719A, G719C, G719D,G719S, S720C, S720F, G721V, W731Stop, P733L, K739-I744 (insertion),V742I, V742A, E746-A750 (deletion), E746K, L747S, L747-E749 (deletion),L747-T751 (deletion), L747-P753 (deletion), G746-S752 (deletion), T751I,S752Y, K754 (deletion), S752-I759 (deletion), A750P, D761-E762 (e.g.,residues EAFQ (SEQ ID NO: 1893) insertion), D761N, D761Y, A763V, V765A,A767-S768 (e.g., residues TLA insertion), A767-V769 (e.g., residues ASVinsertion), S768I, S768T, V769L, V769M, V769-D770 (e.g., residue Yinsertion), 770-771 (e.g., residues GL insertion), 770-771 (e.g.,residue G insertion), 770-771 (e.g., residues CV insertion), 770-771(e.g., residues SVD insertion), P772R, 773-774 (e.g., residues NPHinsertion), H773R, H773L, V774M, 774-775 (e.g., residues HV insertion),R776H, R776C, G779F, T783A, T784F, T854A, V786L, T790M, L792P, P794H,L798F, R803W, H805R, D807H, G810S, N826S, Y827 (nonsense), R831H, R832C,R832H, L833F, L833V, H835L, D837V, L838M, L838P, A839V, N842H, V843L,T847K, T847I, H850N, V851A, I853T, F856L, L858R, L858M, L861Q, L861R,G863D, Q894L, G917A, E967A, D1006Y, P1019L, S1042N, R1100S, H1129Y,T1141S, S1153I, Q1164R, L1167M, or a combination thereof of the EGFRpolypeptide.

In some instances, the polynucleic acid molecule hybridizes to a targetregion of EGFR DNA or RNA comprising one or more mutations. In someembodiments, the polynucleic acid molecule hybridizes to a target regionof EGFR DNA or RNA comprising one or more mutations in exon 18, exon 19,exon 20, exon 21, exon 22, or a combination thereof.

In some embodiments, a polynucleic acid molecule hybridizes to a targetregion of EGFR DNA or RNA comprising one or more mutations at positionscorresponding to amino acid residues 34, 38, 45, 62, 63, 77, 78, 108,114, 120, 140, 148, 149, 160, 177, 178, 189, 191, 198, 220, 222, 223,229, 237, 240, 244, 252, 254, 255, 256, 263, 270, 273, 276, 282, 288,289, 301, 303, 304, 309, 314, 326, 331, 354, 363, 373, 337, 380, 384,393, 427, 428, 437, 441, 447, 465, 475, 515, 526, 527, 531, 536, 541,546, 571, 588, 589, 596, 596, 598, 602, 614, 620, 628, 636, 641, 645,651, 671, 689, 694, 700, 709, 712, 714, 715, 716, 719, 720, 721, 731,733, 739-744, 742, 746-750, 746-752, 746, 747, 747-749, 747-751,747-753, 751, 752, 754, 752-759, 750, 761-762, 761, 763, 765, 767-768,767-769, 768, 769, 769-770, 770-771, 772, 773-774, 773, 774, 774-775,776, 779, 783, 784, 786, 790, 792, 794, 798, 803, 805, 807, 810, 826,827, 831, 832, 833, 835, 837, 838, 839, 842, 843, 847, 850, 851, 853,854, 856, 858, 861, 863, 894, 917, 967, 1006, 1019, 1042, 1100, 1129,1141, 1153, 1164, 1167, or a combination thereof of the EGFRpolypeptide. In some embodiments, the polynucleic acid moleculehybridizes to a target region of EGFR DNA or RNA comprising one or moremutations at positions corresponding to amino acid residues 747, 761,790, 854, 858, or a combination thereof of the EGFR polypeptide. In someembodiments, the polynucleic acid molecule hybridizes to a target regionof EGFR DNA or RNA comprising one or more mutations at positionscorresponding to amino acid residues 761, 790, 858, or a combinationthereof of the EGFR polypeptide. In some embodiments, the polynucleicacid molecule hybridizes to a target region of EGFR DNA or RNAcomprising a mutation at a position corresponding to amino acid residue747 of the EGFR polypeptide. In some embodiments, the polynucleic acidmolecule hybridizes to a target region of EGFR DNA or RNA comprising amutation at a position corresponding to amino acid residue 761 of theEGFR polypeptide. In some embodiments, the polynucleic acid moleculehybridizes to a target region of EGFR DNA or RNA comprising a mutationat a position corresponding to amino acid residue 790 of the EGFRpolypeptide. In some embodiments, the polynucleic acid moleculehybridizes to a target region of EGFR DNA or RNA comprising a mutationat a position corresponding to amino acid residue 854 of the EGFRpolypeptide. In some embodiments, the polynucleic acid moleculehybridizes to a target region of EGFR DNA or RNA comprising a mutationat a position corresponding to amino acid residue 858 of the EGFRpolypeptide.

In some embodiments, a polynucleic acid molecule hybridizes to a targetregion of EGFR DNA or RNA comprising one or more mutations selected fromT34M, L38V, E45Q, L62R, G63R, G63K, S77F, F78L, R108K, R108G, E114K,A120P, L140V, V148M, R149W, E160K, S177P, M178I, K189T, D191N, S198R,S220P, R222L, R222C, S223Y, S229C, A237Y, C240Y, R244G, R252C, R252P,F254I, R255 (nonsense mutation), D256Y, T263P, Y270C, T273A, Q276(nonsense), E282K, G288 (frame shift), A289D, A289V, A289T, A289N,A289D, V301 (deletion), D303H, H304Y, R309Q, D314N, C326R, G331R, T354M,T363I, P373Q, R337S, S380 (frame shift), T384S, D393Y, R427L, G428S,S437Y, V441I, S447Y, G465R, I475V, C515S, C526S, R527L, R531 (nonsense),V536M, L541I, P546Q, C571S, G588S, P589L, P596L, P596S, P596R, P596L,G598V, G598A, E602G, G614D, C620Y, C620W, C628Y, C628F, C636Y, T638M,P641H, S645C, V651M, R671C, V689M, P694S, N700D, E709A, E709K, E709Q,E709K, F712L, K714N, I715S, K716R, G719A, G719C, G719D, G719S, S720C,S720F, G721V, W731Stop, P733L, K739-I744 (insertion), V742I, V742A,E746-A750 (deletion), E746K, L747S, L747-E749 (deletion), L747-T751(deletion), L747-P753 (deletion), G746-S752 (deletion), T751I, S752Y,K754 (deletion), S752-I759 (deletion), A750P, D761-E762 (e.g., residuesEAFQ (SEQ ID NO: 1893) insertion), D761N, D761Y, A763V, V765A, A767-S768(e.g., residues TLA insertion), A767-V769 (e.g., residues ASVinsertion), S768I, S768T, V769L, V769M, V769-D770 (e.g., residue Yinsertion), 770-771 (e.g., residues GL insertion), 770-771 (e.g.,residue G insertion), 770-771 (e.g., residues CV insertion), 770-771(e.g., residues SVD insertion), P772R, 773-774 (e.g., residues NPHinsertion), H773R, H773L, V774M, 774-775 (e.g., residues HV insertion),R776H, R776C, G779F, T783A, T784F, T854A, V786L, T790M, L792P, P794H,L798F, R803W, H805R, D807H, G810S, N826S, Y827 (nonsense), R831H, R832C,R832H, L833F, L833V, H835L, D837V, L838M, L838P, A839V, N842H, V843L,T847K, T847I, H850N, V851A, I853T, F856L, L858R, L858M, L861Q, L861R,G863D, Q894L, G917A, E967A, D1006Y, P1019L, S1042N, R1100S, H1129Y,T1141S, S1153I, Q1164R, L1167M, or a combination thereof of the EGFRpolypeptide. In some embodiments, the polynucleic acid moleculehybridizes to a target region of EGFR DNA or RNA comprising one or moremutations selected from L747S, D761Y, T790M, T854A, L858R, or acombination thereof of the EGFR polypeptide. In some embodiments, thepolynucleic acid molecule hybridizes to a target region of EGFR DNA orRNA comprising one or more mutations selected from D761Y, T790M, L858R,or a combination thereof of the EGFR polypeptide. In some embodiments,the polynucleic acid molecule hybridizes to a target region of EGFR DNAor RNA comprising mutation L747S of the EGFR polypeptide. In someembodiments, the polynucleic acid molecule hybridizes to a target regionof EGFR DNA or RNA comprising mutation D761Y of the EGFR polypeptide. Insome embodiments, the polynucleic acid molecule hybridizes to a targetregion of EGFR DNA or RNA comprising mutation T790M of the EGFRpolypeptide. In some embodiments, the polynucleic acid moleculehybridizes to a target region of EGFR DNA or RNA comprising mutationT854A of the EGFR polypeptide. In some embodiments, the polynucleic acidmolecule hybridizes to a target region of EGFR DNA or RNA comprisingmutation L858R of the EGFR polypeptide.

In some embodiments, a polynucleic acid molecule comprises a sequencethat hybridizes to a target sequence illustrated in Table 1. In someembodiments, the polynucleic acid molecule hybridizes to an EGFR targetsequence selected from SEQ ID NOs: 1-376. In some cases, the polynucleicacid molecule hybridizes to an EGFR target sequence selected from SEQ IDNOs: 1-376 with less than 5 mismatched bases, with less than 4mismatched bases, with less than 3 mismatched bases, with less than 2mismatched bases, or with 1 mismatched base. In some cases, thepolynucleic acid molecule hybridizes to an EGFR target sequence selectedfrom SEQ ID NOs: 1-376 with less than 4 mismatched bases.

In some embodiments, a polynucleic acid molecule comprises a sequencehaving at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%,97%, 98%, 99%, or 100% sequence identity to a sequence listed in Table2, Table 3, or Table 6. In some embodiments, the polynucleic acidmolecule comprises a sequence having at least 50%, 55%, 60%, 65%, 70%,75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identityto SEQ ID NOs: 377-1892. In some embodiments, the polynucleic acidmolecule comprises a sequence having at least 50% sequence identity toSEQ ID NOs: 377-1892. In some embodiments, the polynucleic acid moleculecomprises a sequence having at least 60% sequence identity to SEQ IDNOs: 377-1892. In some embodiments, the polynucleic acid moleculecomprises a sequence having at least 70% sequence identity to SEQ IDNOs: 377-1892. In some embodiments, the polynucleic acid moleculecomprises a sequence having at least 75% sequence identity to SEQ IDNOs: 377-1892. In some embodiments, the polynucleic acid moleculecomprises a sequence having at least 80% sequence identity to SEQ IDNOs: 377-1892. In some embodiments, the polynucleic acid moleculecomprises a sequence having at least 85% sequence identity to SEQ IDNOs: 377-1892. In some embodiments, the polynucleic acid moleculecomprises a sequence having at least 90% sequence identity to SEQ IDNOs: 377-1892. In some embodiments, the polynucleic acid moleculecomprises a sequence having at least 95% sequence identity to SEQ IDNOs: 377-1892. In some embodiments, the polynucleic acid moleculecomprises a sequence having at least 96% sequence identity to SEQ IDNOs: 377-1892. In some embodiments, the polynucleic acid moleculecomprises a sequence having at least 97% sequence identity to SEQ IDNOs: 377-1892. In some embodiments, the polynucleic acid moleculecomprises a sequence having at least 98% sequence identity to SEQ IDNOs: 377-1892. In some embodiments, the polynucleic acid moleculecomprises a sequence having at least 99% sequence identity to SEQ IDNOs: 377-1892. In some embodiments, the polynucleic acid moleculeconsists of SEQ ID NOs: 377-1892.

In some embodiments, a polynucleic acid molecule comprises a firstpolynucleotide and a second polynucleotide. In some instances, the firstpolynucleotide comprises a sequence having at least 50%, 55%, 60%, 65%,70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequenceidentity to SEQ ID NOs: 377-1892. In some cases, the secondpolynucleotide comprises a sequence having at least 50%, 55%, 60%, 65%,70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequenceidentity to SEQ ID NOs: 377-1892. In some cases, the polynucleic acidmolecule comprises a first polynucleotide having at least 50%, 55%, 60%,65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequenceidentity to SEQ ID NOs: 377-1892 and a second polynucleotide having atleast 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%,99%, or 100% sequence identity to SEQ ID NOs: 377-1892.

In some embodiments, a polynucleic acid molecule described hereincomprises RNA or DNA. In some cases, the polynucleic acid moleculecomprises RNA. In some instances, RNA comprises short interfering RNA(siRNA), short hairpin RNA (shRNA), microRNA (miRNA), double-strandedRNA (dsRNA), transfer RNA (tRNA), ribosomal RNA (rRNA), or heterogeneousnuclear RNA (hnRNA). In some instances, RNA comprises shRNA. In someinstances, RNA comprises miRNA. In some instances, RNA comprises dsRNA.In some instances, RNA comprises tRNA. In some instances, RNA comprisesrRNA. In some instances, RNA comprises hnRNA. In some instances, the RNAcomprises siRNA. In some instances, the polynucleic acid moleculecomprises siRNA.

In some embodiments, a polynucleic acid molecule described herein isfrom about 10 to about 50 nucleotides in length. In some instances, thepolynucleic acid molecule is from about 10 to about 30, from about 15 toabout 30, from about 18 to about 25, from about 18 to about 24, fromabout 19 to about 23, or from about 20 to about 22 nucleotides inlength.

In some embodiments, a polynucleic acid molecule is about 50 nucleotidesin length. In some instances, the polynucleic acid molecule is about 45nucleotides in length. In some instances, the polynucleic acid moleculeis about 40 nucleotides in length. In some instances, the polynucleicacid molecule is about 35 nucleotides in length. In some instances, thepolynucleic acid molecule is about 30 nucleotides in length. In someinstances, the polynucleic acid molecule is about 25 nucleotides inlength. In some instances, the polynucleic acid molecule is about 20nucleotides in length. In some instances, the polynucleic acid moleculeis about 19 nucleotides in length. In some instances, the polynucleicacid molecule is about 18 nucleotides in length. In some instances, thepolynucleic acid molecule is about 17 nucleotides in length. In someinstances, the polynucleic acid molecule is about 16 nucleotides inlength. In some instances, the polynucleic acid molecule is about 15nucleotides in length. In some instances, the polynucleic acid moleculeis about 14 nucleotides in length. In some instances, the polynucleicacid molecule is about 13 nucleotides in length. In some instances, thepolynucleic acid molecule is about 12 nucleotides in length. In someinstances, the polynucleic acid molecule is about 11 nucleotides inlength. In some instances, the polynucleic acid molecule is about 10nucleotides in length. In some instances, the polynucleic acid moleculeis between about 10 and about 50 nucleotides in length. In someinstances, the polynucleic acid molecule is from about 10 to about 45nucleotides in length. In some instances, the polynucleic acid moleculeis from about 10 to about 40 nucleotides in length. In some instances,the polynucleic acid molecule is from about 10 to about 35 nucleotidesin length. In some instances, the polynucleic acid molecule is fromabout 10 to about 30 nucleotides in length. In some instances, thepolynucleic acid molecule is from about 10 to about 25 nucleotides inlength. In some instances, the polynucleic acid molecule is from about10 to about 20 nucleotides in length. In some instances, the polynucleicacid molecule is from about 15 to about 25 nucleotides in length. Insome instances, the polynucleic acid molecule is from about 15 to about30 nucleotides in length. In some instances, the polynucleic acidmolecule is from about 12 to about 30 nucleotides in length.

In some embodiments, a polynucleic acid molecule described hereincomprises a first polynucleotide. In some instances, the polynucleicacid molecule comprises a second polynucleotide. In some instances, thepolynucleic acid molecule comprises a first polynucleotide and a secondpolynucleotide. In some instances, the first polynucleotide is a sensestrand or passenger strand. In some instances, the second polynucleotideis an antisense strand or guide strand.

In some embodiments, a polynucleic acid molecule is a firstpolynucleotide. In some embodiments, the first polynucleotide is fromabout 10 to about 50 nucleotides in length. In some instances, the firstpolynucleotide is from about 10 to about 30, from about 15 to about 30,from about 18 to about 25, from about 18 to about 24, from about 19 toabout 23, or from about 20 to about 22 nucleotides in length.

In some instances, a first polynucleotide is about 50 nucleotides inlength. In some instances, the first polynucleotide is about 45nucleotides in length. In some instances, the first polynucleotide isabout 40 nucleotides in length. In some instances, the firstpolynucleotide is about 35 nucleotides in length. In some instances, thefirst polynucleotide is about 30 nucleotides in length. In someinstances, the first polynucleotide is about 25 nucleotides in length.In some instances, the first polynucleotide is about 20 nucleotides inlength. In some instances, the first polynucleotide is about 19nucleotides in length. In some instances, the first polynucleotide isabout 18 nucleotides in length. In some instances, the firstpolynucleotide is about 17 nucleotides in length. In some instances, thefirst polynucleotide is about 16 nucleotides in length. In someinstances, the first polynucleotide is about 15 nucleotides in length.In some instances, the first polynucleotide is about 14 nucleotides inlength. In some instances, the first polynucleotide is about 13nucleotides in length. In some instances, the first polynucleotide isabout 12 nucleotides in length. In some instances, the firstpolynucleotide is about 11 nucleotides in length. In some instances, thefirst polynucleotide is about 10 nucleotides in length. In someinstances, the first polynucleotide is from about 10 to about 50nucleotides in length. In some instances, the first polynucleotide isfrom about 10 to about 45 nucleotides in length. In some instances, thefirst polynucleotide is from about 10 to about 40 nucleotides in length.In some instances, the first polynucleotide is from about 10 to about 35nucleotides in length. In some instances, the first polynucleotide isfrom about 10 to about 30 nucleotides in length. In some instances, thefirst polynucleotide is from about 10 to about 25 nucleotides in length.In some instances, the first polynucleotide is from about 10 to about 20nucleotides in length. In some instances, the first polynucleotide isfrom about 15 to about 25 nucleotides in length. In some instances, thefirst polynucleotide is from about 15 to about 30 nucleotides in length.In some instances, the first polynucleotide is from about 12 to about 30nucleotides in length.

In some embodiments, a polynucleic acid molecule is a secondpolynucleotide. In some embodiments, the second polynucleotide is fromabout 10 to about 50 nucleotides in length. In some instances, thesecond polynucleotide is from about 10 to about 30, from about 15 toabout 30, from about 18 to about 25, from about 18 to about 24, fromabout 19 to about 23, or from about 20 to about 22 nucleotides inlength.

In some instances, a second polynucleotide is about 50 nucleotides inlength. In some instances, the second polynucleotide is about 45nucleotides in length. In some instances, the second polynucleotide isabout 40 nucleotides in length. In some instances, the secondpolynucleotide is about 35 nucleotides in length. In some instances, thesecond polynucleotide is about 30 nucleotides in length. In someinstances, the second polynucleotide is about 25 nucleotides in length.In some instances, the second polynucleotide is about 20 nucleotides inlength. In some instances, the second polynucleotide is about 19nucleotides in length. In some instances, the second polynucleotide isabout 18 nucleotides in length. In some instances, the secondpolynucleotide is about 17 nucleotides in length. In some instances, thesecond polynucleotide is about 16 nucleotides in length. In someinstances, the second polynucleotide is about 15 nucleotides in length.In some instances, the second polynucleotide is about 14 nucleotides inlength. In some instances, the second polynucleotide is about 13nucleotides in length. In some instances, the second polynucleotide isabout 12 nucleotides in length. In some instances, the secondpolynucleotide is about 11 nucleotides in length. In some instances, thesecond polynucleotide is about 10 nucleotides in length. In someinstances, the second polynucleotide is from about 10 to about 50nucleotides in length. In some instances, the second polynucleotide isfrom about 10 to about 45 nucleotides in length. In some instances, thesecond polynucleotide is from about 10 to about 40 nucleotides inlength. In some instances, the second polynucleotide is from about 10 toabout 35 nucleotides in length. In some instances, the secondpolynucleotide is from about 10 to about 30 nucleotides in length. Insome instances, the second polynucleotide is from about 10 to about 25nucleotides in length. In some instances, the second polynucleotide isfrom about 10 to about 20 nucleotides in length. In some instances, thesecond polynucleotide is from about 15 to about 25 nucleotides inlength. In some instances, the second polynucleotide is from about 15 toabout 30 nucleotides in length. In some instances, the secondpolynucleotide is from about 12 to about 30 nucleotides in length.

In some embodiments, a polynucleic acid molecule comprises a firstpolynucleotide and a second polynucleotide. In some instances, thepolynucleic acid molecule further comprises a blunt terminus, anoverhang, or a combination thereof. In some instances, the bluntterminus is a 5′ blunt terminus, a 3′ blunt terminus, or both. In somecases, the overhang is a 5′ overhang, 3′ overhang, or both. In somecases, the overhang comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 non-basepairing nucleotides. In some cases, the overhang comprises 1, 2, 3, 4,5, or 6 non-base pairing nucleotides. In some cases, the overhangcomprises 1, 2, 3, or 4 non-base pairing nucleotides. In some cases, theoverhang comprises 1 non-base pairing nucleotide. In some cases, theoverhang comprises 2 non-base pairing nucleotides. In some cases, theoverhang comprises 3 non-base pairing nucleotides. In some cases, theoverhang comprises 4 non-base pairing nucleotides.

In some embodiments, the sequence of a polynucleic acid molecule is atleast 40%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98%, 99%,or 99.5% complementary to a target sequence described herein. In someembodiments, the sequence of the polynucleic acid molecule is at least50% complementary to a target sequence described herein. In someembodiments, the sequence of the polynucleic acid molecule is at least60% complementary to a target sequence described herein. In someembodiments, the sequence of the polynucleic acid molecule is at least70% complementary to a target sequence described herein. In someembodiments, the sequence of the polynucleic acid molecule is at least80% complementary to a target sequence described herein. In someembodiments, the sequence of the polynucleic acid molecule is at least90% complementary to a target sequence described herein. In someembodiments, the sequence of the polynucleic acid molecule is at least95% complementary to a target sequence described herein. In someembodiments, the sequence of the polynucleic acid molecule is at least99% complementary to a target sequence described herein. In someinstances, the sequence of the polynucleic acid molecule is 100%complementary to a target sequence described herein.

In some embodiments, the sequence of a polynucleic acid molecule has 5or less mismatches to a target sequence described herein. In someembodiments, the sequence of the polynucleic acid molecule has 4 or lessmismatches to a target sequence described herein. In some instances, thesequence of the polynucleic acid molecule has 3 or less mismatches to atarget sequence described herein. In some cases, the sequence of thepolynucleic acid molecule has 2 or less mismatches to a target sequencedescribed herein. In some cases, the sequence of the polynucleic acidmolecule has 1 or less mismatches to a target sequence described herein.

In some embodiments, the specificity of a polynucleic acid molecule thathybridizes to a target sequence described herein is a 95%, 98%, 99%,99.5% or 100% sequence complementarity of the polynucleic acid moleculeto a target sequence. In some instances, the hybridization is a highstringent hybridization condition.

In some embodiments, the polynucleic acid molecule hybridizes to atleast 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or morecontiguous bases of a target sequence described herein. In someembodiments, the polynucleic acid molecule hybridizes to at least 8contiguous bases of a target sequence described herein. In someembodiments, the polynucleic acid molecule hybridizes to at least 9contiguous bases of a target sequence described herein. In someembodiments, the polynucleic acid molecule hybridizes to at least 10contiguous bases of a target sequence described herein. In someembodiments, the polynucleic acid molecule hybridizes to at least 11contiguous bases of a target sequence described herein. In someembodiments, the polynucleic acid molecule hybridizes to at least 12contiguous bases of a target sequence described herein. In someembodiments, the polynucleic acid molecule hybridizes to at least 15contiguous bases of a target sequence described herein. In someembodiments, the polynucleic acid molecule hybridizes to at least 18contiguous bases of a target sequence described herein.

In some embodiments, a polynucleic acid molecule has reduced off-targeteffect. In some instances, “off-target” or “off-target effects” refer toany instance in which a polynucleic acid polymer directed against agiven target causes an unintended effect by interacting either directlyor indirectly with another mRNA sequence, a DNA sequence or a cellularprotein or other moiety. In some instances, an “off-target effect”occurs when there is a simultaneous degradation of other transcripts dueto partial homology or complementarity between that other transcript andthe sense and/or antisense strand of the polynucleic acid molecule.

In some embodiments, a polynucleic acid molecule comprises natural,synthetic, or artificial nucleotide analogues or bases. In some cases,the polynucleic acid molecule comprises combinations of DNA, RNA and/ornucleotide analogues. In some instances, the synthetic or artificialnucleotide analogues or bases comprise modifications at one or more ofribose moiety, phosphate moiety, nucleoside moiety, or a combinationthereof.

In some embodiments, nucleotide analogues or artificial nucleotide basecomprise a nucleic acid with a modification at a 2′ hydroxyl group ofthe ribose moiety. In some instances, the modification includes an H,OR, R, halo, SH, SR, NH2, NHR, NR2, or CN, wherein R is an alkyl moiety.Exemplary alkyl moiety includes, but is not limited to, halogens,sulfurs, thiols, thioethers, thioesters, amines (primary, secondary, ortertiary), amides, ethers, esters, alcohols and oxygen. In someinstances, the alkyl moiety further comprises a modification. In someinstances, the modification comprises an azo group, a keto group, analdehyde group, a carboxyl group, a nitro group, a nitroso, group, anitrile group, a heterocycle (e.g., imidazole, hydrazino orhydroxylamino) group, an isocyanate or cyanate group, or a sulfurcontaining group (e.g., sulfoxide, sulfone, sulfide, or disulfide). Insome instances, the alkyl moiety further comprises a heterosubstitution. In some instances, the carbon of the heterocyclic group issubstituted by a nitrogen, oxygen or sulfur. In some instances, theheterocyclic substitution includes but is not limited to, morpholino,imidazole, and pyrrolidino.

In some instances, the modification at the 2′ hydroxyl group is a2′-O-methyl modification or a 2′-O-methoxyethyl (2′-O-MOE) modification.In some cases, the 2′-O-methyl modification adds a methyl group to the2′ hydroxyl group of the ribose moiety whereas the 2′O-methoxyethylmodification adds a methoxyethyl group to the 2′ hydroxyl group of theribose moiety. Exemplary chemical structures of a 2′-O-methylmodification of an adenosine molecule and 2′O-methoxyethyl modificationof a uridine are illustrated below.

In some instances, the modification at the 2′ hydroxyl group is a2′-O-aminopropyl modification in which an extended amine groupcomprising a propyl linker binds the amine group to the 2′ oxygen. Insome instances, this modification neutralizes the phosphate-derivedoverall negative charge of the oligonucleotide molecule by introducingone positive charge from the amine group per sugar and thereby improvescellular uptake properties due to its zwitterionic properties. Anexemplary chemical structure of a 2′-O-aminopropyl nucleosidephosphoramidite is illustrated below.

In some instances, the modification at the 2′ hydroxyl group is a lockedor bridged ribose modification (e.g., locked nucleic acid or LNA) inwhich the oxygen molecule bound at the 2′ carbon is linked to the 4′carbon by a methylene group, thus forming a2′-C,4′-C-oxy-methylene-linked bicyclic ribonucleotide monomer.Exemplary representations of the chemical structure of LNA areillustrated below. The representation shown to the left highlights thechemical connectivities of an LNA monomer. The representation shown tothe right highlights the locked 3′-endo (³E) conformation of thefuranose ring of an LNA monomer.

In some instances, the modification at the 2′ hydroxyl group comprisesethylene nucleic acids (ENA) such as for example 2′-4′-ethylene-bridgednucleic acid, which locks the sugar conformation into a C₃′-endo sugarpuckering conformation. ENA are part of the bridged nucleic acids classof modified nucleic acids that also comprises LNA. Exemplary chemicalstructures of the ENA and bridged nucleic acids are illustrated below.

In some embodiments, additional modifications at the 2′ hydroxyl groupinclude 2′-deoxy, T-deoxy-2′-fluoro, 2′-O-aminopropyl (2′-O-AP),2′-O-dimethylaminoethyl (2′-O-DMAOE), 2′-O-dimethylaminopropyl(2′-O-DMAP), T-O-dimethylaminoethyloxyethyl (2′-O-DMAEOE), or2′-O—N-methylacetamido (2′-O-NMA).

In some embodiments, nucleotide analogues comprise modified bases suchas, but not limited to, 5-propynyluridine, 5-propynylcytidine,6-methyladenine, 6-methylguanine, N,N,-dimethyladenine, 2-propyladenine,2-propylguanine, 2-aminoadenine, 1-methylinosine, 3-methyluridine,5-methylcytidine, 5-methyluridine and other nucleotides having amodification at the 5 position, 5-(2-amino) propyl uridine,5-halocytidine, 5-halouridine, 4-acetylcytidine, 1-methyladenosine,2-methyladenosine, 3-methylcytidine, 6-methyluridine, 2-methylguanosine,7-methylguanosine, 2, 2-dimethylguanosine, 5-methylaminoethyluridine,5-methyloxyuridine, deazanucleotides (such as 7-deaza-adenosine,6-azouridine, 6-azocytidine, or 6-azothymidine), 5-methyl-2-thiouridine,other thio bases (such as 2-thiouridine, 4-thiouridine, and2-thiocytidine), dihydrouridine, pseudouridine, queuosine, archaeosine,naphthyl and substituted naphthyl groups, any O-and N-alkylated purinesand pyrimidines (such as N6-methyladenosine,5-methylcarbonylmethyluridine, uridine 5-oxyacetic acid, pyridine-4-one,or pyridine-2-one), phenyl and modified phenyl groups (such asaminophenol or 2,4,6-trimethoxy benzene), modified cytosines that act asG-clamp nucleotides, 8-substituted adenines and guanines, 5-substituteduracils and thymines, azapyrimidines, carboxyhydroxyalkyl nucleotides,carboxyalkylaminoalkyi nucleotides, and alkylcarbonylalkylatednucleotides. Modified nucleotides also include those nucleotides thatare modified with respect to the sugar moiety, as well as nucleotideshaving sugars or analogs thereof that are not ribosyl. For example, thesugar moieties, in some cases, are or are based on mannoses, arabinoses,glucopyranoses, galactopyranoses, 4′-thioribose, and other sugars,heterocycles, or carbocycles. The term nucleotide also includes what areknown in the art as universal bases. By way of example, universal basesinclude, but are not limited to, 3-nitropyrrole, 5-nitroindole, ornebularine.

In some embodiments, nucleotide analogues further comprise morpholinos,peptide nucleic acids (PNAs), methylphosphonate nucleotides,thiolphosphonate nucleotides, 2′-fluoro N3-P5′-phosphoramidites,1′,5′-anhydrohexitol nucleic acids (HNAs), or a combination thereof.Morpholino or phosphorodiamidate morpholino oligo (PMO) comprisessynthetic molecules whose structure mimics natural nucleic acidstructure but deviates from the normal sugar and phosphate structures.In some instances, the five member ribose ring is substituted with a sixmember morpholino ring containing four carbons, one nitrogen, and oneoxygen. In some cases, the ribose monomers are linked by aphosphordiamidate group instead of a phosphate group. In such cases, thebackbone alterations remove all positive and negative charges makingmorpholinos neutral molecules capable of crossing cellular membraneswithout the aid of cellular delivery agents such as those used bycharged oligonucleotides.

In some embodiments, peptide nucleic acid (PNA) does not contain sugarring or phosphate linkage and the bases are attached and appropriatelyspaced by oligoglycine-like molecules, therefore eliminating a backbonecharge.

In some embodiments, one or more modifications optionally occur at theinternucleotide linkage. In some instances, modified internucleotidelinkage includes, but is not limited to, phosphorothioates;phosphorodithioates; methylphosphonates; 5′-alkylenephosphonates;5′-methylphosphonate; 3′-alkylene phosphonates; borontrifluoridates;borano phosphate esters and selenophosphates of 3′-5′linkage or2′-5′linkage; phosphotriesters; thionoalkylphosphotriesters; hydrogenphosphonate linkages; alkyl phosphonates; alkylphosphonothioates;arylphosphonothioates; phosphoroselenoates; phosphorodiselenoates;phosphinates; phosphoramidates; 3′-alkylphosphoramidates;aminoalkylphosphoramidates; thionophosphoramidates;phosphoropiperazidates; phosphoroanilothioates; phosphoroanilidates;ketones; sulfones; sulfonamides; carbonates; carbamates;methylenehydrazos; methylenedimethylhydrazos; formacetals;thioformacetals; oximes; methyleneiminos; methylenemethyliminos;thioamidates; linkages with riboacetyl groups; aminoethyl glycine; silylor siloxane linkages; alkyl or cycloalkyl linkages with or withoutheteroatoms of, for example, 1 to 10 carbons that are saturated orunsaturated and/or substituted and/or contain heteroatoms; linkages withmorpholino structures, amides, or polyamides wherein the bases areattached to the aza nitrogens of the backbone directly or indirectly;and combinations thereof.

In some instances, the modification is a methyl or thiol modificationsuch as methylphosphonate or thiolphosphonate modification. Exemplarythiolphosphonate nucleotide (left) and methylphosphonate nucleotide(right) are illustrated below.

In some instances, a modified nucleotide includes, but is not limitedto, 2′-fluoro N3-P5′-phosphoramidites illustrated as:

In some instances, a modified nucleotide includes, but is not limitedto, hexitol nucleic acid (or 1′,5′-anhydrohexitol nucleic acids (HNA))illustrated as:

In some embodiments, one or more modifications further optionallyinclude modifications of the ribose moiety, phosphate backbone and thenucleoside, or modifications of the nucleotide analogues at the 3′ orthe 5′ terminus. For example, the 3′ terminus optionally include a 3′cationic group, or by inverting the nucleoside at the 3′-terminus with a3′-3′ linkage. In another alternative, the 3′-terminus is optionallyconjugated with an aminoalkyl group, e.g., a 3′ C5-aminoalkyl dT. In anadditional alternative, the 3′-terminus is optionally conjugated with anabasic site, e.g., with an apurinic or apyrimidinic site. In someinstances, the 5′-terminus is conjugated with an aminoalkyl group, e.g.,a 5′-O-alkylamino substituent. In some cases, the 5′-terminus isconjugated with an abasic site, e.g., with an apurinic or apyrimidinicsite.

In some embodiments, a polynucleic acid molecule comprises one or moreartificial nucleotide analogues described herein. In some instances, thepolynucleic acid molecule comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,12, 13, 14, 15, 16, 17, 18, 20, 25, or more artificial nucleotideanalogues described herein. In some embodiments, the artificialnucleotide analogues include 2′-O-methyl, 2′-O-methoxyethyl (2′-O-MOE),2′-O-aminopropyl, 2′-deoxy, T-deoxy-2′-fluoro, 2′-O-aminopropyl(2′-O-AP), 2′-O-dimethylaminoethyl (2′-O-DMAOE),2′-O-dimethylaminopropyl (2′-O-DMAP), T-O-dimethylaminoethyloxyethyl(2′-O-DMAEOE), or 2′-O—N-methylacetamido (2′-O-NMA) modified, LNA, ENA,PNA, HNA, morpholino, methylphosphonate nucleotides, thiolphosphonatenucleotides, 2′-fluoro N3-P5′-phosphoramidites, or a combinationthereof. In some instances, the polynucleic acid molecule comprises 1,2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 20, 25, ormore of the artificial nucleotide analogues selected from 2′-O-methyl,2′-O-methoxyethyl (2′-O-MOE), 2′-O-aminopropyl, 2′-deoxy,T-deoxy-2′-fluoro, 2′-O-aminopropyl (2′-O-AP), 2′-O-dimethylaminoethyl(2′-O-DMAOE), 2′-O-dimethylaminopropyl (2′-O-DMAP),T-O-dimethylaminoethyloxyethyl (2′-O-DMAEOE), or 2′-O—N-methylacetamido(2′-O-NMA) modified, LNA, ENA, PNA, HNA, morpholino, methylphosphonatenucleotides, thiolphosphonate nucleotides, 2′-fluoroN3-P5′-phosphoramidites, or a combination thereof. In some instances,the polynucleic acid molecule comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, 15, 16, 17, 18, 20, 25, or more of 2′-O-methyl modifiednucleotides. In some instances, the polynucleic acid molecule comprises1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 20, 25,or more of 2′-O-methoxyethyl (2′-O-MOE) modified nucleotides. In someinstances, the polynucleic acid molecule comprises 1, 2, 3, 4, 5, 6, 7,8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 20, 25, or more ofthiolphosphonate nucleotides.

In some instances, a polynucleic acid molecule comprises at least oneof: from about 5% to about 100% modification, from about 10% to about100% modification, from about 20% to about 100% modification, from about30% to about 100% modification, from about 40% to about 100%modification, from about 50% to about 100% modification, from about 60%to about 100% modification, from about 70% to about 100% modification,from about 80% to about 100% modification, and from about 90% to about100% modification. In some instances, the polynucleic acid molecule is apolynucleic acid molecule of SEQ ID NOs: 377-1128.

In some cases, a polynucleic acid molecule comprises at least one of:from about 10% to about 90% modification, from about 20% to about 90%modification, from about 30% to about 90% modification, from about 40%to about 90% modification, from about 50% to about 90% modification,from about 60% to about 90% modification, from about 70% to about 90%modification, and from about 80% to about 100% modification. In someinstances, the polynucleic acid molecule is a polynucleic acid moleculeof SEQ ID NOs: 377-1128.

In some cases, a polynucleic acid molecule comprises at least one of:from about 10% to about 80% modification, from about 20% to about 80%modification, from about 30% to about 80% modification, from about 40%to about 80% modification, from about 50% to about 80% modification,from about 60% to about 80% modification, and from about 70% to about80% modification. In some instances, the polynucleic acid molecule is apolynucleic acid molecule of SEQ ID NOs: 377-1128.

In some instances, a polynucleic acid molecule comprises at least oneof: from about 10% to about 70% modification, from about 20% to about70% modification, from about 30% to about 70% modification, from about40% to about 70% modification, from about 50% to about 70% modification,and from about 60% to about 70% modification. In some instances, thepolynucleic acid molecule is a polynucleic acid molecule of SEQ ID NOs:377-1128.

In some instances, a polynucleic acid molecule comprises at least oneof: from about 10% to about 60% modification, from about 20% to about60% modification, from about 30% to about 60% modification, from about40% to about 60% modification, and from about 50% to about 60%modification. In some instances, the polynucleic acid molecule is apolynucleic acid molecule of SEQ ID NOs: 377-1128.

In some cases, a polynucleic acid molecule comprises at least one of:from about 10% to about 50% modification, from about 20% to about 50%modification, from about 30% to about 50% modification, and from about40% to about 50% modification. In some instances, the polynucleic acidmolecule is a polynucleic acid molecule of SEQ ID NOs: 377-1128.

In some cases, a polynucleic acid molecule comprises at least one of:from about 10% to about 40% modification, from about 20% to about 40%modification, and from about 30% to about 40% modification. In someinstances, the polynucleic acid molecule is a polynucleic acid moleculeof SEQ ID NOs: 377-1128.

In some cases, a polynucleic acid molecule comprises at least one of:from about 10% to about 30% modification, and from about 20% to about30% modification. In some instances, the polynucleic acid molecule is apolynucleic acid molecule of SEQ ID NOs: 377-1128.

In some cases, a polynucleic acid molecule comprises from about 10% toabout 20% modification. In some instances, the polynucleic acid moleculeis a polynucleic acid molecule of SEQ ID NOs: 377-1128.

In some cases, a polynucleic acid molecule comprises from about 15% toabout 90%, from about 20% to about 80%, from about 30% to about 70%, orfrom about 40% to about 60% modifications. In some instances, thepolynucleic acid molecule is a polynucleic acid molecule of SEQ ID NOs:377-1128.

In additional cases, a polynucleic acid molecule comprises at leastabout 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99%modification. In some instances, the polynucleic acid molecule is apolynucleic acid molecule of SEQ ID NOs: 377-1128.

In some embodiments, a polynucleic acid molecule comprises at leastabout 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8,about 9, about 10, about 11, about 12, about 13, about 14, about 15,about 16, about 17, about 18, about 19, about 20, about 21, about 22, ormore modifications. In some instances, the polynucleic acid molecule isa polynucleic acid molecule of SEQ ID NOs: 377-1128.

In some instances, a polynucleic acid molecule comprises at least about1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about9, about 10, about 11, about 12, about 13, about 14, about 15, about 16,about 17, about 18, about 19, about 20, about 21, about 22, or moremodified nucleotides. In some instances, the polynucleic acid moleculeis a polynucleic acid molecule of SEQ ID NOs: 377-1128.

In some instances, from about 5 to about 100% of a polynucleic acidmolecule comprise an artificial nucleotide analogue described herein. Insome instances, about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%,55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% of the polynucleicacid molecule comprise an artificial nucleotide analogue describedherein. In some instances, about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%,45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% of apolynucleic acid molecule of SEQ ID NOs: 377-1892 comprise an artificialnucleotide analogue described herein. In some instances, about 5%, 10%,15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%,85%, 90%, 95%, or 100% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 5% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 10% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 15% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 20% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 25% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 30% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 35% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 40% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 45% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 50% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 55% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 60% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 65% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 70% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 75% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 80% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 85% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 90% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 95% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 96% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 97% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 98% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 99% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome instances, about 100% of a polynucleic acid molecule of SEQ ID NOs:377-1128 comprise an artificial nucleotide analogue described herein. Insome embodiments, the artificial nucleotide analogue comprises2′-O-methyl, 2′-O-methoxyethyl (2′-O-MOE), 2′-O-aminopropyl, 2′-deoxy,T-deoxy-2′-fluoro, 2′-O-aminopropyl (2′-O-AP), 2′-O-dimethylaminoethyl(2′-O-DMAOE), 2′-O-dimethylaminopropyl (2′-O-DMAP),T-O-dimethylaminoethyloxyethyl (2′-O-DMAEOE), or 2′-O—N-methylacetamido(2′-O-NMA) modified, LNA, ENA, PNA, HNA, morpholino, methylphosphonatenucleotides, thiolphosphonate nucleotides, 2′-fluoroN3-P5′-phosphoramidites, or a combination thereof.

In some embodiments, a polynucleic acid molecule comprises from about 1to about 25 modifications in which the modification comprises anartificial nucleotide analogues described herein. In some embodiments, apolynucleic acid molecule of SEQ ID NOs: 377-1128 comprises from about 1to about 25 modifications in which the modifications comprise anartificial nucleotide analogue described herein. In some embodiments, apolynucleic acid molecule of SEQ ID NOs: 377-1128 comprises about 1modification in which the modification comprises an artificialnucleotide analogue described herein. In some embodiments, a polynucleicacid molecule of SEQ ID NOs: 377-1128 comprises about 2 modifications inwhich the modifications comprise an artificial nucleotide analoguedescribed herein. In some embodiments, a polynucleic acid molecule ofSEQ ID NOs: 377-1128 comprises about 3 modifications in which themodifications comprise an artificial nucleotide analogue describedherein. In some embodiments, a polynucleic acid molecule of SEQ ID NOs:377-1128 comprises about 4 modifications in which the modificationscomprise an artificial nucleotide analogue described herein. In someembodiments, a polynucleic acid molecule of SEQ ID NOs: 377-1128comprises about 5 modifications in which the modifications comprise anartificial nucleotide analogue described herein. In some embodiments, apolynucleic acid molecule of SEQ ID NOs: 377-1128 comprises about 6modifications in which the modifications comprise an artificialnucleotide analogue described herein. In some embodiments, a polynucleicacid molecule of SEQ ID NOs: 377-1128 comprises about 7 modifications inwhich the modifications comprise an artificial nucleotide analoguedescribed herein. In some embodiments, a polynucleic acid molecule ofSEQ ID NOs: 377-1128 comprises about 8 modifications in which themodifications comprise an artificial nucleotide analogue describedherein. In some embodiments, a polynucleic acid molecule of SEQ ID NOs:377-1128 comprises about 9 modifications in which the modificationscomprise an artificial nucleotide analogue described herein. In someembodiments, a polynucleic acid molecule of SEQ ID NOs: 377-1128comprises about 10 modifications in which the modifications comprise anartificial nucleotide analogue described herein. In some embodiments, apolynucleic acid molecule of SEQ ID NOs: 377-1128 comprises about 11modifications in which the modifications comprise an artificialnucleotide analogue described herein. In some embodiments, a polynucleicacid molecule of SEQ ID NOs: 377-1128 comprises about 12 modificationsin which the modifications comprise an artificial nucleotide analoguedescribed herein. In some embodiments, a polynucleic acid molecule ofSEQ ID NOs: 377-1128 comprises about 13 modifications in which themodifications comprise an artificial nucleotide analogue describedherein. In some embodiments, a polynucleic acid molecule of SEQ ID NOs:377-1128 comprises about 14 modifications in which the modificationscomprise an artificial nucleotide analogue described herein. In someembodiments, a polynucleic acid molecule of SEQ ID NOs: 377-1128comprises about 15 modifications in which the modifications comprise anartificial nucleotide analogue described herein. In some embodiments, apolynucleic acid molecule of SEQ ID NOs: 377-1128 comprises about 16modifications in which the modifications comprise an artificialnucleotide analogue described herein. In some embodiments, a polynucleicacid molecule of SEQ ID NOs: 377-1128 comprises about 17 modificationsin which the modifications comprise an artificial nucleotide analoguedescribed herein. In some embodiments, a polynucleic acid molecule ofSEQ ID NOs: 377-1128 comprises about 18 modifications in which themodifications comprise an artificial nucleotide analogue describedherein. In some embodiments, a polynucleic acid molecule of SEQ ID NOs:377-1128 comprises about 19 modifications in which the modificationscomprise an artificial nucleotide analogue described herein. In someembodiments, a polynucleic acid molecule of SEQ ID NOs: 377-1128comprises about 20 modifications in which the modifications comprise anartificial nucleotide analogue described herein. In some embodiments, apolynucleic acid molecule of SEQ ID NOs: 377-1128 comprises about 21modifications in which the modifications comprise an artificialnucleotide analogue described herein. In some embodiments, a polynucleicacid molecule of SEQ ID NOs: 377-1128 comprises about 22 modificationsin which the modifications comprise an artificial nucleotide analoguedescribed herein. In some embodiments, a polynucleic acid molecule ofSEQ ID NOs: 377-1128 comprises about 23 modifications in which themodifications comprise an artificial nucleotide analogue describedherein. In some embodiments, a polynucleic acid molecule of SEQ ID NOs:377-1128 comprises about 24 modifications in which the modificationscomprise an artificial nucleotide analogue described herein. In someembodiments, a polynucleic acid molecule of SEQ ID NOs: 377-1128comprises about 25 modifications in which the modifications comprise anartificial nucleotide analogue described herein.

In some instances, a polynucleic acid molecule that comprises anartificial nucleotide analogue comprises a sequence selected from SEQ IDNOs: 1129-1892.

In some embodiments, a polynucleic acid molecule is assembled from twoseparate polynucleotides wherein one polynucleotide comprises the sensestrand and the second polynucleotide comprises the antisense strand ofthe polynucleic acid molecule. In other embodiments, the sense strand isconnected to the antisense strand via a linker molecule, which in someinstances, is a polynucleotide linker or a non-nucleotide linker.

In some embodiments, a polynucleic acid molecule comprises a sensestrand and antisense strand, wherein pyrimidine nucleotides in the sensestrand comprise 2′-O-methylpyrimidine nucleotides and purine nucleotidesin the sense strand comprise 2′-deoxy purine nucleotides. In someembodiments, a polynucleic acid molecule comprises a sense strand andantisense strand, wherein pyrimidine nucleotides present in the sensestrand comprise 2′-deoxy-2′-fluoro pyrimidine nucleotides and whereinpurine nucleotides present in the sense strand comprise 2′-deoxy purinenucleotides.

In some embodiments, a polynucleic acid molecule comprises a sensestrand and antisense strand, wherein the pyrimidine nucleotides whenpresent in said antisense strand are 2′-deoxy-2′-fluoro pyrimidinenucleotides and the purine nucleotides when present in said antisensestrand are 2′-O-methyl purine nucleotides.

In some embodiments, a polynucleic acid molecule comprises a sensestrand and antisense strand, wherein the pyrimidine nucleotides whenpresent in said antisense strand are 2′-deoxy-2′-fluoro pyrimidinenucleotides and wherein the purine nucleotides when present in saidantisense strand comprise 2′-deoxy-purine nucleotides.

In some embodiments, a polynucleic acid molecule comprises a sensestrand and antisense strand, wherein the sense strand includes aterminal cap moiety at the 5′-end, the 3′-end, or both of the 5′ and 3′ends of the sense strand. In other embodiments, the terminal cap moietyis an inverted deoxy abasic moiety.

In some embodiments, a polynucleic acid molecule comprises a sensestrand and an antisense strand, wherein the antisense strand comprises aphosphate backbone modification at the 3′ end of the antisense strand.In some instances, the phosphate backbone modification is aphosphorothioate.

In some embodiments, a polynucleic acid molecule comprises a sensestrand and an antisense strand, wherein the antisense strand comprises aglyceryl modification at the 3′ end of the antisense strand.

In some embodiments, a polynucleic acid molecule comprises a sensestrand and an antisense strand, in which the sense strand comprises oneor more (for example, about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,14, 15, 16, 17, 18, 19, 20, or more) phosphorothioate internucleotidelinkages, and/or one or more (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10or more) 2′-deoxy, 2′-O-methyl, 2′-deoxy-2′-fluoro, and/or about one ormore (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more) universal basemodified nucleotides, and optionally a terminal cap molecule at the3′-end, the 5′-end, or both of the 3′- and 5′-ends of the sense strand;and in which the antisense strand comprises about 1 to about 10 or more,specifically about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15,16, 17, 18, 19, 20, or more, phosphorothioate internucleotide linkages,and/or one or more (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more)2′-deoxy, 2′-O-methyl, 2′-deoxy-2′-fluoro, and/or one or more (e.g.,about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more) universal base modifiednucleotides, and optionally a terminal cap molecule at the 3′-end, the5′-end, or both of the 3′- and 5′-ends of the antisense strand. In otherembodiments, one or more (for example about 1, 2, 3, 4, 5, 6, 7, 8, 9,10, or more) pyrimidine nucleotides of the sense and/or antisense strandare chemically-modified with 2′-deoxy, 2′-O-methyl and/or2′-deoxy-2′-fluoro nucleotides, with or without one or more (for exampleabout 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more) phosphorothioateinternucleotide linkages and/or a terminal cap molecule at the 3′-end,the 5′-end, or both of the 3′- and 5′-ends, being present in the same ordifferent strand.

In some embodiments, a polynucleic acid molecule comprises a sensestrand and an antisense strand, in which the sense strand comprisesabout 1 to about 25 (for example, about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) phosphorothioateinternucleotide linkages, and/or one or more (e.g., about 1, 2, 3, 4, 5,6, 7, 8, 9, 10, or more) 2′-deoxy, 2′-O-methyl, 2′-deoxy-2′-fluoro,and/or one or more (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more)universal base modified nucleotides, and optionally a terminal capmolecule at the 3-end, the 5′-end, or both of the 3′- and 5′-ends of thesense strand; and in which the antisense strand comprises about 1 toabout 25 or more (for example about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,12, 13, 14, 15, 16, 17, 18, 19, 20, or more) phosphorothioateinternucleotide linkages, and/or one or more (e.g., about 1, 2, 3, 4, 5,6, 7, 8, 9, 10 or more) 2′-deoxy, 2′-O-methyl, 2′-deoxy-2′-fluoro,and/or one or more (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more)universal base modified nucleotides, and optionally a terminal capmolecule at the 3′-end, the 5′-end, or both of the 3′- and 5′-ends ofthe antisense strand. In other embodiments, one or more (for exampleabout 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more) pyrimidine nucleotides ofthe sense and/or antisense strand are chemically-modified with 2′-deoxy,2′-O-methyl and/or 2′-deoxy-2′-fluoro nucleotides, with or without about1 to about 25 or more (for example about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) phosphorothioateinternucleotide linkages and/or a terminal cap molecule at the 3′-end,the 5′-end, or both of the 3′- and 5′-ends, being present in the same ordifferent strand.

In some embodiments, a polynucleic acid molecule comprises a sensestrand and an antisense strand, in which the antisense strand comprisesone or more (for example, about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12,13, 14, 15, 16, 17, 18, 19, 20, or more) phosphorothioateinternucleotide linkages, and/or about one or more (e.g., about 1, 2, 3,4, 5, 6, 7, 8, 9, 10 or more) 2′-deoxy, 2′-O-methyl, 2′-deoxy-2′-fluoro,and/or one or more (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more)universal base modified nucleotides, and optionally a terminal capmolecule at the 3′-end, the 5′-end, or both of the 3′- and 5′-ends ofthe sense strand; and wherein the antisense strand comprises about 1 toabout 10 or more, specifically about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 ormore, phosphorothioate internucleotide linkages, and/or one or more(e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more) 2′-deoxy,2′-O-methyl, 2′-deoxy-2′-fluoro, and/or one or more (e.g., about 1, 2,3, 4, 5, 6, 7, 8, 9, 10 or more) universal base modified nucleotides,and optionally a terminal cap molecule at the 3′-end, the 5′-end, orboth of the 3′- and 5′-ends of the antisense strand. In otherembodiments, one or more (for example about 1, 2, 3, 4, 5, 6, 7, 8, 9,10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) pyrimidinenucleotides of the sense and/or antisense strand are chemically-modifiedwith 2′-deoxy, 2′-O-methyl and/or 2′-deoxy-2′-fluoro nucleotides, withor without one or more (for example, about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10or more) phosphorothioate internucleotide linkages and/or a terminal capmolecule at the 3′-end, the 5′-end, or both of the 3′ and 5′-ends, beingpresent in the same or different strand.

In some embodiments, a polynucleic acid molecule comprises a sensestrand and an antisense strand, in which the antisense strand comprisesabout 1 to about 25 or more (for example, about 1, 2, 3, 4, 5, 6, 7, 8,9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or more) phosphorothioateinternucleotide linkages, and/or one or more (e.g., about 1, 2, 3, 4, 5,6, 7, 8, 9, 10 or more) 2′-deoxy, 2′-O-methyl, 2′-deoxy-2′-fluoro,and/or one or more (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more)universal base modified nucleotides, and optionally a terminal capmolecule at the 3′-end, the 5′-end, or both of the 3′- and 5′-ends ofthe sense strand; and wherein the antisense strand comprises about 1 toabout 25 or more (for example about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,12, 13, 14, 15, 16, 17, 18, 19, 20, or more) phosphorothioateinternucleotide linkages, and/or one or more (e.g., about 1, 2, 3, 4, 5,6, 7, 8, 9, 10 or more) 2′-deoxy, 2′-O-methyl, 2′-deoxy-2′-fluoro,and/or one or more (e.g., about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more)universal base modified nucleotides, and optionally a terminal capmolecule at the 3′-end, the 5′-end, or both of the 3′- and 5′-ends ofthe antisense strand. In other embodiments, one or more (for exampleabout 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more) pyrimidine nucleotides ofthe sense and/or antisense strand are chemically-modified with 2′-deoxy,2′-O-methyl and/or 2′-deoxy-2′-fluoro nucleotides, with or without about1 to about 5 (for example about 1, 2, 3, 4, 5 or more) phosphorothioateinternucleotide linkages and/or a terminal cap molecule at the 3′-end,the 5′-end, or both of the 3′- and 5′-ends, being present in the same ordifferent strand.

In some embodiments, a polynucleic acid molecule described herein is achemically-modified short interfering nucleic acid molecule having about1 to about 25 (for example, about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12,13, 14, 15, 16, 17, 18, 19, 20 or more) phosphorothioate internucleotidelinkages in each strand of the polynucleic acid molecule.

In another embodiment, a polynucleic acid molecule described hereincomprises 2′-5′ internucleotide linkages. In some instances, the 2′-5′internucleotide linkage(s) is at the 3′-end, the 5′-end, or both of the3′- and 5′-ends of one or both sequence strands. In addition instances,the 2′-5′ internucleotide linkage(s) is present at various otherpositions within one or both sequence strands (for example, about 1, 2,3, 4, 5, 6, 7, 8, 9, 10, or more) including every internucleotidelinkage of a pyrimidine nucleotide in one or both strands of thepolynucleic acid molecule comprise a 2′-5′ internucleotide linkage, orabout 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more, including everyinternucleotide linkage of a purine nucleotide in one or both strands ofthe polynucleic acid molecule comprise a 2′-5′ internucleotide linkage.

In some embodiments, a polynucleic acid molecule is a single strandedpolynucleic acid molecule that mediates RNAi activity in a cell orreconstituted in vitro system, wherein the polynucleic acid moleculecomprises a single stranded polynucleotide having complementarity to atarget nucleic acid sequence, and wherein one or more pyrimidinenucleotides present in the polynucleic acid are 2′-deoxy-2′-fluoropyrimidine nucleotides (e.g., wherein all pyrimidine nucleotides are2′-deoxy-2′-fluoro pyrimidine nucleotides or alternately a plurality ofpyrimidine nucleotides are 2′-deoxy-2′-fluoro pyrimidine nucleotides),and wherein one or more purine nucleotides present in the polynucleicacid are 2′-deoxy purine nucleotides (e.g., wherein all purinenucleotides are 2′-deoxy purine nucleotides or alternately a pluralityof purine nucleotides are 2′-deoxy purine nucleotides), and a terminalcap modification, that is optionally present at the 3′-end, the 5′-end,or both of the 3′ and 5′-ends of the antisense sequence, the polynucleicacid molecule optionally further comprising about 1 to about 4 (e.g.,about 1, 2, 3, or 4) terminal 2′-deoxynucleotides at the 3′-end of thepolynucleic acid molecule, wherein the terminal nucleotides furthercomprise one or more (e.g., 1, 2, 3, or 4) phosphorothioateinternucleotide linkages, and wherein the polynucleic acid moleculeoptionally further comprises a terminal phosphate group, such as a5′-terminal phosphate group.

In some cases, one or more artificial nucleotide analogues describedherein are resistant toward nucleases such as for example ribonucleasesuch as RNase H, deoxyribunuclease such as DNase, or exonuclease such as5′-3′ exonuclease and 3′-5′ exonuclease, when compared to naturalpolynucleic acid molecules. In some instances, artificial nucleotideanalogues comprising 2′-O-methyl, 2′-O-methoxyethyl (2′-O-MOE),2′-O-aminopropyl, 2′-deoxy, T-deoxy-2′-fluoro, 2′-O-aminopropyl(2′-O-AP), 2′-O-dimethylaminoethyl (2′-O-DMAOE),2′-O-dimethylaminopropyl (2′-O-DMAP), T-O-dimethylaminoethyloxyethyl(2′-O-DMAEOE), or 2′-O—N-methylacetamido (2′-O-NMA) modified, LNA, ENA,PNA, HNA, morpholino, methylphosphonate nucleotides, thiolphosphonatenucleotides, 2′-fluoro N3-P5′-phosphoramidites, or combinations thereofare resistant toward nucleases such as for example ribonuclease such asRNase H, deoxyribunuclease such as DNase, or exonuclease such as 5′-3′exonuclease and 3′-5′ exonuclease. In some instances, 2′-O-methylmodified polynucleic acid molecule is nuclease resistant (e.g., RNase H,DNase, 5′-3′ exonuclease or 3′-5′ exonuclease resistant). In someinstances, 2′O-methoxyethyl (2′-O-MOE) modified polynucleic acidmolecule is nuclease resistant (e.g., RNase H, DNase, 5′-3′ exonucleaseor 3′-5′ exonuclease resistant). In some instances, 2′-O-aminopropylmodified polynucleic acid molecule is nuclease resistant (e.g., RNase H,DNase, 5′-3′ exonuclease or 3′-5′ exonuclease resistant). In someinstances, 2′-deoxy modified polynucleic acid molecule is nucleaseresistant (e.g., RNase H, DNase, 5′-3′ exonuclease or 3′-5′ exonucleaseresistant). In some instances, T-deoxy-2′-fluoro modified polynucleicacid molecule is nuclease resistant (e.g., RNase H, DNase, 5′-3′exonuclease or 3′-5′ exonuclease resistant). In some instances,2′-O-aminopropyl (2′-O-AP) modified polynucleic acid molecule isnuclease resistant (e.g., RNase H, DNase, 5′-3′ exonuclease or 3′-5′exonuclease resistant). In some instances, 2′-O-dimethylaminoethyl(2′-O-DMAOE) modified polynucleic acid molecule is nuclease resistant(e.g., RNase H, DNase, 5′-3′ exonuclease or 3′-5′ exonucleaseresistant). In some instances, 2′-O-dimethylaminopropyl (2′-O-DMAP)modified polynucleic acid molecule is nuclease resistant (e.g., RNase H,DNase, 5′-3′ exonuclease or 3′-5′ exonuclease resistant). In someinstances, T-O-dimethylaminoethyloxyethyl (2′-O-DMAEOE) modifiedpolynucleic acid molecule is nuclease resistant (e.g., RNase H, DNase,5′-3′ exonuclease or 3′-5′ exonuclease resistant). In some instances,2′-O—N-methylacetamido (2′-O-NMA) modified polynucleic acid molecule isnuclease resistant (e.g., RNase H, DNase, 5′-3′ exonuclease or 3′-5′exonuclease resistant). In some instances, LNA modified polynucleic acidmolecule is nuclease resistant (e.g., RNase H, DNase, 5′-3′ exonucleaseor 3′-5′ exonuclease resistant). In some instances, ENA modifiedpolynucleic acid molecule is nuclease resistant (e.g., RNase H, DNase,5′-3′ exonuclease or 3′-5′ exonuclease resistant). In some instances,HNA modified polynucleic acid molecule is nuclease resistant (e.g.,RNase H, DNase, 5′-3′ exonuclease or 3′-5′ exonuclease resistant). Insome instances, morpholinos is nuclease resistant (e.g., RNase H, DNase,5′-3′ exonuclease or 3′-5′ exonuclease resistant). In some instances,PNA modified polynucleic acid molecule is resistant to nucleases (e.g.,RNase H, DNase, 5′-3′ exonuclease or 3′-5′ exonuclease resistant). Insome instances, methylphosphonate nucleotides modified polynucleic acidmolecule is nuclease resistant (e.g., RNase H, DNase, 5′-3′ exonucleaseor 3′-5′ exonuclease resistant). In some instances, thiolphosphonatenucleotides modified polynucleic acid molecule is nuclease resistant(e.g., RNase H, DNase, 5′-3′ exonuclease or 3′-5′ exonucleaseresistant). In some instances, polynucleic acid molecule comprising2′-fluoro N3-P5′-phosphoramidites is nuclease resistant (e.g., RNase H,DNase, 5′-3′ exonuclease or 3′-5′ exonuclease resistant). In someinstances, the 5′ conjugates described herein inhibit 5′-3′exonucleolytic cleavage. In some instances, the 3′ conjugates describedherein inhibit exonucleolytic cleavage.

In some embodiments, one or more of the artificial nucleotide analoguesdescribed herein have increased binding affinity toward their mRNAtarget relative to an equivalent natural polynucleic acid molecule. Theone or more of the artificial nucleotide analogues comprising2′-O-methyl, 2′-O-methoxyethyl (2′-O-MOE), 2′-O-aminopropyl, 2′-deoxy,T-deoxy-2′-fluoro, 2′-O-aminopropyl (2′-O-AP), 2′-O-dimethylaminoethyl(2′-O-DMAOE), 2′-O-dimethylaminopropyl (2′-O-DMAP),T-O-dimethylaminoethyloxyethyl (2′-O-DMAEOE), or 2′-O—N-methylacetamido(2′-O-NMA) modified, LNA, ENA, PNA, HNA, morpholino, methylphosphonatenucleotides, thiolphosphonate nucleotides, or 2′-fluoroN3-P5′-phosphoramidites have increased binding affinity toward theirmRNA target relative to an equivalent natural polynucleic acid molecule.In some instances, 2′-O-methyl-modified polynucleic acid molecule hasincreased binding affinity toward their mRNA target relative to anequivalent natural polynucleic acid molecule. In some instances,2′-O-methoxyethyl (2′-O-MOE) modified polynucleic acid molecule hasincreased binding affinity toward their mRNA target relative to anequivalent natural polynucleic acid molecule. In some instances,2′-O-aminopropyl modified polynucleic acid molecule has increasedbinding affinity toward their mRNA target relative to an equivalentnatural polynucleic acid molecule. In some instances, 2′-deoxy modifiedpolynucleic acid molecule has increased binding affinity toward theirmRNA target relative to an equivalent natural polynucleic acid molecule.In some instances, T-deoxy-2′-fluoro modified polynucleic acid moleculehas increased binding affinity toward their mRNA target relative to anequivalent natural polynucleic acid molecule. In some instances,2′-O-aminopropyl (2′-O-AP) modified polynucleic acid molecule hasincreased binding affinity toward their mRNA target relative to anequivalent natural polynucleic acid molecule. In some instances,2′-O-dimethylaminoethyl (2′-O-DMAOE) modified polynucleic acid moleculehas increased binding affinity toward their mRNA target relative to anequivalent natural polynucleic acid molecule. In some instances,2′-O-dimethylaminopropyl (2′-O-DMAP) modified polynucleic acid moleculehas increased binding affinity toward their mRNA target relative to anequivalent natural polynucleic acid molecule. In some instances,T-O-dimethylaminoethyloxyethyl (2′-O-DMAEOE) modified polynucleic acidmolecule has increased binding affinity toward their mRNA targetrelative to an equivalent natural polynucleic acid molecule. In someinstances, 2′-O—N-methylacetamido (2′-O-NMA) modified polynucleic acidmolecule has increased binding affinity toward their mRNA targetrelative to an equivalent natural polynucleic acid molecule. In someinstances, LNA-modified polynucleic acid molecule has increased bindingaffinity toward their mRNA target relative to an equivalent naturalpolynucleic acid molecule. In some instances, ENA-modified polynucleicacid molecule has increased binding affinity toward their mRNA targetrelative to an equivalent natural polynucleic acid molecule. In someinstances, PNA-modified polynucleic acid molecule has increased bindingaffinity toward their mRNA target relative to an equivalent naturalpolynucleic acid molecule. In some instances, HNA-modified polynucleicacid molecule has increased binding affinity toward their mRNA targetrelative to an equivalent natural polynucleic acid molecule. In someinstances, morpholino-modified polynucleic acid molecule has increasedbinding affinity toward their mRNA target relative to an equivalentnatural polynucleic acid molecule. In some instances, methylphosphonatenucleotide-modified polynucleic acid molecule has increased bindingaffinity toward their mRNA target relative to an equivalent naturalpolynucleic acid molecule. In some instances, thiolphosphonatenucleotide-modified polynucleic acid molecule has increased bindingaffinity toward their mRNA target relative to an equivalent naturalpolynucleic acid molecule. In some instances, polynucleic acid moleculecomprising 2′-fluoro N3-P5′-phosphoramidites has increased bindingaffinity toward their mRNA target relative to an equivalent naturalpolynucleic acid molecule. In some cases, the increased affinity isillustrated with a lower Kd, a higher melt temperature (Tm), or acombination thereof.

In some embodiments, a polynucleic acid molecule described herein is achirally pure (or stereo pure) polynucleic acid molecule, or apolynucleic acid molecule comprising a single enantiomer. In someinstances, the polynucleic acid molecule comprises L-nucleotide. In someinstances, the polynucleic acid molecule comprises D-nucleotides. Insome instance, a polynucleic acid molecule composition comprises lessthan 30%, 25%, 20%, 15%, 10%, 5%, 4%, 3%, 2%, 1%, or less of its mirrorenantiomer. In some cases, a polynucleic acid molecule compositioncomprises less than 30%, 25%, 20%, 15%, 10%, 5%, 4%, 3%, 2%, 1%, or lessof a racemic mixture. In some instances, the polynucleic acid moleculeis a polynucleic acid molecule described in: U.S. Patent PublicationNos: 2014/194610 and 2015/211006; and PCT Publication No.: WO2015107425.

In some embodiments, a polynucleic acid molecule described herein isfurther modified to include an aptamer-conjugating moiety. In someinstances, the aptamer conjugating moiety is a DNA aptamer-conjugatingmoiety. In some instances, the aptamer-conjugating moiety is Alphamer(Centauri Therapeutics), which comprises an aptamer portion thatrecognizes a specific cell-surface target and a portion that presents aspecific epitopes for attaching to circulating antibodies. In someinstance, a polynucleic acid molecule described herein is furthermodified to include an aptamer-conjugating moiety as described in: U.S.Pat. Nos. 8,604,184, 8,591,910, and 7,850,975.

In additional embodiments, a polynucleic acid molecule described hereinis modified to increase its stability. In some embodiment, thepolynucleic acid molecule is RNA (e.g., siRNA), and the polynucleic acidmolecule is modified to increase its stability. In some instances, thepolynucleic acid molecule is modified by one or more of themodifications described above to increase its stability. In some cases,the polynucleic acid molecule is modified at the 2′ hydroxyl position,such as by 2′-O-methyl, 2′-O-methoxyethyl (2′-O-MOE), 2′-O-aminopropyl,2′-deoxy, T-deoxy-2′-fluoro, 2′-O-aminopropyl (2′-O-AP),2′-O-dimethylaminoethyl (2′-O-DMAOE), 2′-O-dimethylaminopropyl(2′-O-DMAP), T-O-dimethylaminoethyloxyethyl (2′-O-DMAEOE), or2′-O—N-methylacetamido (2′-O-NMA) modification or by a locked or bridgedribose conformation (e.g., LNA or ENA). In some cases, the polynucleicacid molecule is modified by 2′-O-methyl and/or 2′-O-methoxyethylribose. In some cases, the polynucleic acid molecule also includesmorpholinos, PNAs, HNA, methylphosphonate nucleotides, thiolphosphonatenucleotides, and/or 2′-fluoro N3-P5′-phosphoramidites to increase itsstability. In some instances, the polynucleic acid molecule is achirally pure (or stereo pure) polynucleic acid molecule. In someinstances, the chirally pure (or stereo pure) polynucleic acid moleculeis modified to increase its stability. Suitable modifications to the RNAto increase stability for delivery will be apparent to the skilledperson.

In some embodiments, a polynucleic acid molecule describe herein hasRNAi activity that modulates expression of RNA encoded by EGFR. In someinstances, a polynucleic acid molecule described herein is adouble-stranded siRNA molecule that down-regulates expression of EGFR,wherein one of the strands of the double-stranded siRNA moleculecomprises a nucleotide sequence that is complementary to a nucleotidesequence of EGFR or RNA encoded by EGFR or a portion thereof, andwherein the second strand of the double-stranded siRNA moleculecomprises a nucleotide sequence substantially similar to the nucleotidesequence of EGFR or RNA encoded by EGFR or a portion thereof. In somecases, a polynucleic acid molecule described herein is a double-strandedsiRNA molecule that down-regulates expression of EGFR, wherein eachstrand of the siRNA molecule comprises about 15 to 25, 18 to 24, or 19to about 23 nucleotides, and wherein each strand comprises at leastabout 14, 17, or 19 nucleotides that are complementary to thenucleotides of the other strand. In some cases, a polynucleic acidmolecule described herein is a double-stranded siRNA molecule thatdown-regulates expression of EGFR, wherein each strand of the siRNAmolecule comprises about 19 to about 23 nucleotides, and wherein eachstrand comprises at least about 19 nucleotides that are complementary tothe nucleotides of the other strand. In some instances, the RNAiactivity occurs within a cell. In other instances, the RNAi activityoccurs in a reconstituted in vitro system.

In some embodiments, a polynucleic acid molecule describe herein hasRNAi activity that modulates expression of RNA encoded by EGFR. In someinstances, a polynucleic acid molecule described herein is asingle-stranded siRNA molecule that down-regulates expression of EGFR,wherein the single-stranded siRNA molecule comprises a nucleotidesequence that is complementary to a nucleotide sequence of EGFR or RNAencoded by EGFR or a portion thereof. In some cases, a polynucleic acidmolecule described herein is a single-stranded siRNA molecule thatdown-regulates expression of EGFR, wherein the siRNA molecule comprisesabout 15 to 25, 18 to 24, or 19 to about 23 nucleotides. In some cases,a polynucleic acid molecule described herein is a single-stranded siRNAmolecule that down-regulates expression of EGFR, wherein the siRNAmolecule comprises about 19 to about 23 nucleotides. In some instances,the RNAi activity occurs within a cell. In other instances, the RNAiactivity occurs in a reconstituted in vitro system.

In some instances, a polynucleic acid molecule is a double-strandedpolynucleotide molecule comprising self-complementary sense andantisense regions, wherein the antisense region comprises a nucleotidesequence that is complementary to a nucleotide sequence in a targetnucleic acid molecule or a portion thereof and the sense region has anucleotide sequence corresponding to the target nucleic acid sequence ora portion thereof. In some instances, the polynucleic acid molecule isassembled from two separate polynucleotides, where one strand is thesense strand and the other is the antisense strand, wherein theantisense and sense strands are self-complementary (e.g., each strandcomprises a nucleotide sequence that is complementary to the nucleotidesequence in the other strand; such as where the antisense strand andsense strand form a duplex or double-stranded structure, for examplewherein the double-stranded region is about 19, 20, 21, 22, 23, or morebase pairs); the antisense strand comprises a nucleotide sequence thatis complementary to a nucleotide sequence in a target nucleic acidmolecule or a portion thereof and the sense strand comprises anucleotide sequence corresponding to the target nucleic acid sequence ora portion thereof. Alternatively, the polynucleic acid molecule isassembled from a single oligonucleotide, where the self-complementarysense and antisense regions of the polynucleic acid molecule are linkedby means of a nucleic acid based or non-nucleic acid-based linker(s).

In some cases, a polynucleic acid molecule is a polynucleotide with aduplex, asymmetric duplex, hairpin, or asymmetric hairpin secondarystructure, having self-complementary sense and antisense regions,wherein the antisense region comprises a nucleotide sequence that iscomplementary to a nucleotide sequence in a separate target nucleic acidmolecule or a portion thereof and the sense region has a nucleotidesequence corresponding to the target nucleic acid sequence or a portionthereof. In other cases, the polynucleic acid molecule is a circularsingle-stranded polynucleotide having two or more loop structures and astem comprising self-complementary sense and antisense regions, whereinthe antisense region comprises a nucleotide sequence that iscomplementary to a nucleotide sequence in a target nucleic acid moleculeor a portion thereof and the sense region has a nucleotide sequencecorresponding to the target nucleic acid sequence or a portion thereof,and wherein the circular polynucleotide is processed either in vivo orin vitro to generate an active polynucleic acid molecule capable ofmediating RNAi. In additional cases, the polynucleic acid molecule alsocomprises a single-stranded polynucleotide having a nucleotide sequencecomplementary to a nucleotide sequence in a target nucleic acid moleculeor a portion thereof (for example, where such polynucleic acid moleculedoes not require the presence within the polynucleic acid molecule of anucleotide sequence corresponding to the target nucleic acid sequence ora portion thereof), wherein the single stranded polynucleotide furthercomprises a terminal phosphate group, such as a 5′-phosphate (see forexample Martinez et al., 2002, Cell., 110, 563-574 and Schwarz et al.,2002, Molecular Cell, 10, 537-568), or 5′,3′-diphosphate.

In some instances, an asymmetric duplex is a linear polynucleic acidmolecule comprising an antisense region, a loop portion that comprisesnucleotides or non-nucleotides, and a sense region that comprises fewernucleotides than the antisense region to the extent that the senseregion has enough complimentary nucleotides to base pair with theantisense region and form a duplex with loop. For example, an asymmetrichairpin polynucleic acid molecule comprises an antisense region havinglength sufficient to mediate RNAi in a cell or in vitro system (e.g.about 19 to about 22 nucleotides) and a loop region comprising about 4to about 8 nucleotides, and a sense region having about 3 to about 18nucleotides that are complementary to the antisense region. In somecases, the asymmetric hairpin polynucleic acid molecule also comprises a5′-terminal phosphate group that is chemically modified. In additionalcases, the loop portion of the asymmetric hairpin polynucleic acidmolecule comprises nucleotides, non-nucleotides, linker molecules, orconjugate molecules.

In some embodiments, an asymmetric duplex is a polynucleic acid moleculehaving two separate strands comprising a sense region and an antisenseregion, wherein the sense region comprises fewer nucleotides than theantisense region to the extent that the sense region has enoughcomplimentary nucleotides to base pair with the antisense region andform a duplex. For example, an asymmetric duplex polynucleic acidmolecule comprises an antisense region having length sufficient tomediate RNAi in a cell or in vitro system (e.g. about 19 to about 22nucleotides) and a sense region having about 3 to about 18 nucleotidesthat are complementary to the antisense region.

In some cases, a universal base refers to nucleotide base analogs thatform base pain with each of the natural DNA/RNA bases with littlediscrimination between them. Non-limiting examples of universal basesinclude C-phenyl, C-naphthyl and other aromatic derivatives, inosine,azole carboxamides, and nitroazole derivatives such as 3-nitropyrrole,4-nitroindole 5-nitroindole, and 6-nitroindole as known in the art (seefor example Loakes, 2001, Nucleic Acids Research, 29, 2437-2447).

Polynucleic Acid Molecule Synthesis

In some embodiments, a polynucleic acid molecule described herein isconstructed using chemical synthesis and/or enzymatic ligation reactionsusing procedures known in the art. For example, a polynucleic acidmolecule is chemically synthesized using naturally occurring nucleotidesor variously modified nucleotides designed to increase the biologicalstability of the molecules or to increase the physical stability of theduplex formed between the polynucleic acid molecule and target nucleicacids. Exemplary methods include those described in: U.S. Pat. Nos.5,142,047; 5,185,444; 5,889,136; 6,008,400; and 6,111,086; PCTPublication No. WO2009099942; or European Publication No. 1579015.Additional exemplary methods include those described in: Griffey et al.,“2′-O-aminopropyl ribonucleotides: a zwitterionic modification thatenhances the exonuclease resistance and biological activity of antisenseoligonucleotides,” J. Med. Chem. 39(26):5100-5109 (1997)); Obika, et al.“Synthesis of 2′-O,4′-C-methyleneuridine and -cytidine. Novel bicyclicnucleosides having a fixed C3, -endo sugar puckering”. TetrahedronLetters 38 (50): 8735 (1997); Koizumi, M. “ENA oligonucleotides astherapeutics”. Current opinion in molecular therapeutics 8 (2): 144-149(2006); and Abramova et al., “Novel oligonucleotide analogues based onmorpholino nucleoside subunits-antisense technologies: new chemicalpossibilities,” Indian Journal of Chemistry 48B:1721-1726 (2009).Alternatively, the polynucleic acid molecule is produced biologicallyusing an expression vector into which a polynucleic acid molecule hasbeen subcloned in an antisense orientation (i.e., RNA transcribed fromthe inserted polynucleic acid molecule will be of an antisenseorientation to a target polynucleic acid molecule of interest).

In some embodiments, a polynucleic acid molecule is synthesized via atandem synthesis methodology, wherein both strands are synthesized as asingle contiguous oligonucleotide fragment or strand separated by acleavable linker which is subsequently cleaved to provide separatefragments or strands that hybridize and permit purification of theduplex.

In some instances, a polynucleic acid molecule is also assembled fromtwo distinct nucleic acid strands or fragments wherein one fragmentincludes the sense region and the second fragment includes the antisenseregion of the molecule.

Additional modification methods for incorporating, for example, sugar,base, and phosphate modifications include: Eckstein et al.,International Publication PCT No. WO 92/07065; Perrault et al. Nature,1990, 344, 565-568; Pieken et al. Science, 1991, 253, 314-317; Usman andCedergren, Trends in Biochem. Sci., 1992, 17, 334-339; Usman et al.International Publication PCT No. WO 93/15187; Sproat, U.S. Pat. No.5,334,711 and Beigelman et al., 1995, J. Biol. Chem., 270, 25702;Beigelman et al., International PCT publication No. WO 97/26270;Beigelman et al., U.S. Pat. No. 5,716,824; Usman et al., U.S. Pat. No.5,627,053; Woolf et al., International PCT Publication No. WO 98/13526;Thompson et al., U.S. Ser. No. 60/082,404 which was filed on Apr. 20,1998; Karpeisky et al., 1998, Tetrahedron Lett., 39, 1131; Earnshaw andGait, 1998, Biopolymers (Nucleic Acid Sciences), 48, 39-55; Verma andEckstein, 1998, Annu. Rev. Biochem., 67, 99-134; and Burlina et al.,1997, Bioorg. Med. Chem., 5, 1999-2010. Such publications describegeneral methods and strategies to determine the location ofincorporation of sugar, base, and/or phosphate modifications and thelike into nucleic acid molecules without modulating catalysis.

In some instances, while chemical modification of the polynucleic acidmolecule internucleotide linkages with phosphorothioate,phosphorodithioate, and/or 5′-methylphosphonate linkages improvesstability, excessive modifications sometimes cause toxicity or decreasedactivity. Therefore, when designing nucleic acid molecules, the amountof these internucleotide linkages in some cases is minimized. In suchcases, the reduction in the concentration of these linkages lowerstoxicity, and increases efficacy and specificity of these molecules.

Diseases

In some embodiments, a polynucleic acid molecule or a pharmaceuticalcomposition described herein is used for the treatment of a disease ordisorder. In some instances, the disease or disorder is a cancer. Insome embodiments, a polynucleic acid molecule or a pharmaceuticalcomposition described herein is used for the treatment of cancer. Insome instances, the cancer is a solid tumor. In some instances, thecancer is a hematologic malignancy. In some instances, the cancer is arelapsed or refractory cancer, or a metastatic cancer. In someinstances, the solid tumor is a relapsed or refractory solid tumor, or ametastatic solid tumor. In some cases, the hematologic malignancy is arelapsed or refractory hematologic malignancy, or a metastatichematologic malignancy.

In some embodiments, the cancer is a solid tumor. Exemplary solid tumorincludes, but is not limited to, anal cancer, appendix cancer, bile ductcancer (i.e., cholangiocarcinoma), bladder cancer, brain tumor, breastcancer, cervical cancer, colon cancer, cancer of Unknown Primary (CUP),esophageal cancer, eye cancer, fallopian tube cancer,gastroenterological cancer, kidney cancer, liver cancer, lung cancer,medulloblastoma, melanoma, oral cancer, ovarian cancer, pancreaticcancer, parathyroid disease, penile cancer, pituitary tumor, prostatecancer, rectal cancer, skin cancer, stomach cancer, testicular cancer,throat cancer, thyroid cancer, uterine cancer, vaginal cancer, or vulvarcancer.

In some instances, a polynucleic acid molecule or a pharmaceuticalcomposition described herein is used for the treatment of a solid tumor.In some instances, a polynucleic acid molecule or a pharmaceuticalcomposition described herein is used for the treatment of anal cancer,appendix cancer, bile duct cancer (i.e., cholangiocarcinoma), bladdercancer, brain tumor, breast cancer, cervical cancer, colon cancer,cancer of Unknown Primary (CUP), esophageal cancer, eye cancer,fallopian tube cancer, gastroenterological cancer, kidney cancer, livercancer, lung cancer, medulloblastoma, melanoma, oral cancer, ovariancancer, pancreatic cancer, parathyroid disease, penile cancer, pituitarytumor, prostate cancer, rectal cancer, skin cancer, stomach cancer,testicular cancer, throat cancer, thyroid cancer, uterine cancer,vaginal cancer, or vulvar cancer. In some instances, the solid tumor isa relapsed or refractory solid tumor, or a metastatic solid tumor.

In some instances, the cancer is a hematologic malignancy. In someinstances, the hematologic malignancy is a leukemia, a lymphoma, amyeloma, a non-Hodgkin's lymphoma, or a Hodgkin's lymphoma. In someinstances, the hematologic malignancy comprises chronic lymphocyticleukemia (CLL), small lymphocytic lymphoma (SLL), high risk CLL, anon-CLL/SLL lymphoma, prolymphocytic leukemia (PLL), follicular lymphoma(FL), diffuse large B-cell lymphoma (DLBCL), mantle cell lymphoma (MCL),Waldenström's macroglobulinemia, multiple myeloma, extranodal marginalzone B cell lymphoma, nodal marginal zone B cell lymphoma, Burkitt'slymphoma, non-Burkitt high grade B cell lymphoma, primary mediastinalB-cell lymphoma (PMBL), immunoblastic large cell lymphoma, precursorB-lymphoblastic lymphoma, B cell prolymphocytic leukemia,lymphoplasmacytic lymphoma, splenic marginal zone lymphoma, plasma cellmyeloma, plasmacytoma, mediastinal (thymic) large B cell lymphoma,intravascular large B cell lymphoma, primary effusion lymphoma, orlymphomatoid granulomatosis.

In some instances, a polynucleic acid molecule or a pharmaceuticalcomposition described herein is used for the treatment of a hematologicmalignancy. In some instances, a polynucleic acid molecule or apharmaceutical composition described herein is used for the treatment ofa leukemia, a lymphoma, a myeloma, a non-Hodgkin's lymphoma, or aHodgkin's lymphoma. In some instances, the hematologic malignancycomprises chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma(SLL), high risk CLL, a non-CLL/SLL lymphoma, prolymphocytic leukemia(PLL), follicular lymphoma (FL), diffuse large B-cell lymphoma (DLBCL),mantle cell lymphoma (MCL), Waldenström's macroglobulinemia, multiplemyeloma, extranodal marginal zone B cell lymphoma, nodal marginal zone Bcell lymphoma, Burkitt's lymphoma, non-Burkitt high grade B celllymphoma, primary mediastinal B-cell lymphoma (PMBL), immunoblasticlarge cell lymphoma, precursor B-lymphoblastic lymphoma, B cellprolymphocytic leukemia, lymphoplasmacytic lymphoma, splenic marginalzone lymphoma, plasma cell myeloma, plasmacytoma, mediastinal (thymic)large B cell lymphoma, intravascular large B cell lymphoma, primaryeffusion lymphoma, or lymphomatoid granulomatosis. In some cases, thehematologic malignancy is a relapsed or refractory hematologicmalignancy, or a metastatic hematologic malignancy.

In some instances, the cancer is an EGFR-associated cancer. In someinstances, a polynucleic acid molecule or a pharmaceutical compositiondescribed herein is used for the treatment of an EGFR-associated cancer.In some instances, the cancer is a solid tumor. In some instances, thecancer is a hematologic malignancy. In some instances, the solid tumoris a relapsed or refractory solid tumor, or a metastatic solid tumor. Insome cases, the hematologic malignancy is a relapsed or refractoryhematologic malignancy, or a metastatic hematologic malignancy. In someinstances, the cancer comprises bladder cancer, breast cancer,colorectal cancer, endometrial cancer, esophageal cancer, glioblastomamultiforme, head and neck cancer, kidney cancer, lung cancer, ovariancancer, pancreatic cancer, prostate cancer, thyroid cancer, acutemyeloid leukemia, CLL, DLBCL, or multiple myeloma.

Pharmaceutical Formulation

In some embodiments, the pharmaceutical formulations described hereinare administered to a subject by multiple administration routesincluding, but not limited to, parenteral (e.g., intravenous,subcutaneous, intramuscular), oral, intranasal, buccal, rectal, ortransdermal administration routes. In some instances, the pharmaceuticalcomposition describe herein is formulated for parenteral (e.g.,intravenous, subcutaneous, intramuscular) administration. In otherinstances, the pharmaceutical composition describe herein is formulatedfor oral administration. In still other instances, the pharmaceuticalcomposition describe herein is formulated for intranasal administration.

In some embodiments, the pharmaceutical formulations include, but arenot limited to, aqueous liquid dispersions, self-emulsifyingdispersions, solid solutions, liposomal dispersions, aerosols, soliddosage forms, powders, immediate release formulations, controlledrelease formulations, fast melt formulations, tablets, capsules, pills,delayed release formulations, extended release formulations, pulsatilerelease formulations, multiparticulate formulations (e.g., nanoparticleformulations), and mixed immediate- and controlled-release formulations.

In some instances, the pharmaceutical formulation includesmultiparticulate formulations. In some instances, the pharmaceuticalformulation includes nanoparticle formulations. In some instances,nanoparticles comprise cMAP, cyclodextrin, or lipids. In some cases,nanoparticles comprise solid lipid nanoparticles, polymericnanoparticles, self-emulsifying nanoparticles, liposomes,microemulsions, or micellar solutions. Additional exemplarynanoparticles include, but are not limited to, paramagneticnanoparticles, superparamagnetic nanoparticles, metal nanoparticles,fullerene-like materials, inorganic nanotubes, dendrimers (such as withcovalently attached metal chelates), nanofibers, nanohorns, nano-onions,nanorods, nanoropes, and quantum dots. In some instances, a nanoparticleis a metal nanoparticle, e.g., a nanoparticle of scandium, titanium,vanadium, chromium, manganese, iron, cobalt, nickel, copper, zinc,yttrium, zirconium, niobium, molybdenum, ruthenium, rhodium, palladium,silver, cadmium, hafnium, tantalum, tungsten, rhenium, osmium, iridium,platinum, gold, gadolinium, aluminum, gallium, indium, tin, thallium,lead, bismuth, magnesium, calcium, strontium, barium, lithium, sodium,potassium, boron, silicon, phosphorus, germanium, arsenic, antimony, andcombinations, alloys, or oxides thereof.

In some instances, a nanoparticle includes a core or a core and a shell,as in a core-shell nanoparticle.

In some instances, a nanoparticle is further coated with molecules forattachment of functional elements (e.g., with one or more of apolynucleic acid molecule or binding moiety described herein). In someinstances, a coating comprises chondroitin sulfate, dextran sulfate,carboxymethyl dextran, alginic acid, pectin, carragheenan, fucoidan,agaropectin, porphyran, karaya gum, gellan gum, xanthan gum, hyaluronicacids, glucosamine, galactosamine, chitin (or chitosan), polyglutamicacid, polyaspartic acid, lysozyme, cytochrome C, ribonuclease,trypsinogen, chymotrypsinogen, α-chymotrypsin, polylysine, polyarginine,histone, protamine, ovalbumin, dextrin, or cyclodextrin. In someinstances, a nanoparticle comprises a graphene-coated nanoparticle.

In some cases, a nanoparticle has at least one dimension of less thanabout 500 nm, 400 nm, 300 nm, 200 nm, or 100 nm.

In some instances, the nanoparticle formulation comprises paramagneticnanoparticles, superparamagnetic nanoparticles, metal nanoparticles,fullerene-like materials, inorganic nanotubes, dendrimers (such as withcovalently attached metal chelates), nanofibers, nanohorns, nano-onions,nanorods, nanoropes or quantum dots. In some instances, a polynucleicacid molecule or a binding moiety described herein is conjugated eitherdirectly or indirectly to the nanoparticle. In some instances, at least1, 5, 10, 15, 20, 30, 40, 50, 60, 70, 80, 90, 100 or more polynucleicacid molecules or binding moieties described herein are conjugatedeither directly or indirectly to a nanoparticle.

In some embodiments, the pharmaceutical formulation comprise a deliveryvector, e.g., a recombinant vector, for the delivery of the polynucleicacid molecule into cells. In some instances, the recombinant vector isDNA plasmid. In other instances, the recombinant vector is a viralvector. Exemplary viral vectors include vectors derived fromadeno-associated virus, retrovirus, adenovirus, or alphavirus. In someinstances, the recombinant vectors capable of expressing the polynucleicacid molecules provide stable expression in target cells. In additionalinstances, viral vectors are used that provide for transient expressionof polynucleic acid molecules.

In some embodiments, the pharmaceutical formulations include a carrieror carrier materials selected on the basis of compatibility with thecomposition disclosed herein, and the release profile properties of thedesired dosage form. Exemplary carrier materials include, e.g., binders,suspending agents, disintegration agents, filling agents, surfactants,solubilizers, stabilizers, lubricants, wetting agents, diluents, and thelike. Pharmaceutically compatible carrier materials include, but are notlimited to, acacia, gelatin, colloidal silicon dioxide, calciumglycerophosphate, calcium lactate, maltodextrin, glycerine, magnesiumsilicate, polyvinylpyrrollidone (PVP), cholesterol, cholesterol esters,sodium caseinate, soy lecithin, taurocholic acid, phosphotidylcholine,sodium chloride, tricalcium phosphate, dipotassium phosphate, celluloseand cellulose conjugates, sugars sodium stearoyl lactylate, carrageenan,monoglyceride, diglyceride, pregelatinized starch, and the like. See,e.g., Remington: The Science and Practice of Pharmacy, Nineteenth Ed(Easton, Pa.: Mack Publishing Company, 1995); Hoover, John E.,Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, Pa.1975; Liberman, H. A. and Lachman, L., Eds., Pharmaceutical DosageForms, Marcel Decker, New York, N.Y., 1980; and Pharmaceutical DosageForms and Drug Delivery Systems, Seventh Ed. (Lippincott Williams &Wilkins 1999).

In some instances, the pharmaceutical formulations further includepH-adjusting agents or buffering agents which include acids such asacetic, boric, citric, lactic, phosphoric and hydrochloric acids; basessuch as sodium hydroxide, sodium phosphate, sodium borate, sodiumcitrate, sodium acetate, sodium lactate andtris-hydroxymethylaminomethane; and buffers such as citrate/dextrose,sodium bicarbonate, and ammonium chloride. Such acids, bases and buffersare included in an amount required to maintain pH of the composition inan acceptable range.

In some instances, the pharmaceutical formulation includes one or moresalts in an amount required to bring osmolality of the composition intoan acceptable range. Such salts include those having sodium, potassiumor ammonium cations and chloride, citrate, ascorbate, borate, phosphate,bicarbonate, sulfate, thiosulfate or bisulfite anions; suitable saltsinclude sodium chloride, potassium chloride, sodium thiosulfate, sodiumbisulfite and ammonium sulfate.

In some instances, the pharmaceutical formulations further includediluent which are used to stabilize compounds because they provide amore stable environment. Salts dissolved in buffered solutions (whichalso provide pH control or maintenance) are utilized as diluents in theart, including, but not limited to a phosphate-buffered saline solution.In certain instances, diluents increase bulk of the composition tofacilitate compression or create sufficient bulk for homogenous blendfor capsule filling. Such compounds include e.g., lactose, starch,mannitol, sorbitol, dextrose, microcrystalline cellulose such asAvicel®; dibasic calcium phosphate, dicalcium phosphate dihydrate;tricalcium phosphate, calcium phosphate; anhydrous lactose, spray-driedlactose; pregelatinized starch, compressible sugar, such as Di-Pac®(Amstar); mannitol, hydroxypropylmethylcellulose,hydroxypropylmethylcellulose acetate stearate, sucrose-based diluents,confectioner's sugar; monobasic calcium sulfate monohydrate, calciumsulfate dihydrate; calcium lactate trihydrate, dextrates; hydrolyzedcereal solids, amylose; powdered cellulose, calcium carbonate; glycine,kaolin; mannitol, sodium chloride; inositol, bentonite, and the like.

In some cases, the pharmaceutical formulations include disintegrationagents or disintegrants to facilitate the breakup or disintegration of asubstance. The term “disintegrate” includes both the dissolution anddispersion of the dosage form when contacted with gastrointestinalfluid. Examples of disintegration agents include a starch, e.g., anatural starch such as corn starch or potato starch, a pregelatinizedstarch such as National 1551 or Amijel®, or sodium starch glycolate suchas Promogel® or Explotab®; a cellulose such as a wood product,methylcrystalline cellulose, e.g., Avicel®, Avicel® PH101, Avicel®PH102, Avicel® PH105, Elcema® P100, Emcocel®, Vivacel®, Ming Tia®, andSolka-Floc®, methylcellulose, croscarmellose, or a cross-linkedcellulose, such as cross-linked sodium carboxymethylcellulose(Ac-Di-Sol®), cross-linked carboxymethylcellulose, or cross-linkedcroscarmellose; a cross-linked starch such as sodium starch glycolate, across-linked polymer such as crospovidone; a cross-linkedpolyvinylpyrrolidone, alginate such as alginic acid or a salt of alginicacid such as sodium alginate, a clay such as Veegum® HV (magnesiumaluminum silicate); a gum such as agar, guar, locust bean, Karaya,pectin, or tragacanth; sodium starch glycolate; bentonite; a naturalsponge; a surfactant; a resin such as a cation-exchange resin; citruspulp; sodium lauryl sulfate; sodium lauryl sulfate in combinationstarch; and the like.

In some instances, the pharmaceutical formulations include fillingagents such as lactose, calcium carbonate, calcium phosphate, dibasiccalcium phosphate, calcium sulfate, microcrystalline cellulose,cellulose powder, dextrose, dextrates, dextran, starches, pregelatinizedstarch, sucrose, xylitol, lactitol, mannitol, sorbitol, sodium chloride,polyethylene glycol, and the like.

Lubricants and glidants are also optionally included in thepharmaceutical formulations described herein for preventing, reducing,or inhibiting adhesion or friction of materials. Exemplary lubricantsinclude, e.g., stearic acid, calcium hydroxide, talc, sodium stearylfumerate, a hydrocarbon such as mineral oil, or hydrogenated vegetableoil such as hydrogenated soybean oil (Sterotex®), higher fatty acids andtheir alkali-metal and alkaline earth metal salts, such as aluminum,calcium, magnesium, zinc, stearic acid, sodium stearates, glycerol,talc, waxes, Stearowet®, boric acid, sodium benzoate, sodium acetate,sodium chloride, leucine, a polyethylene glycol (e.g., PEG-4000) or amethoxypolyethylene glycol such as Carbowax™ sodium oleate, sodiumbenzoate, glyceryl behenate, polyethylene glycol, magnesium or sodiumlauryl sulfate, colloidal silica such as Syloid™, Cab-O-Sil®, a starchsuch as corn starch, silicone oil, a surfactant, and the like.

Plasticizers include compounds used to soften the microencapsulationmaterial or film coatings to make them less brittle. Suitableplasticizers include, e.g., polyethylene glycols such as PEG 300, PEG400, PEG 600, PEG 1450, PEG 3350, and PEG 800, stearic acid, propyleneglycol, oleic acid, triethyl cellulose and triacetin. Plasticizers alsofunction as dispersing agents or wetting agents.

Solubilizers include compounds such as triacetin, triethylcitrate, ethyloleate, ethyl caprylate, sodium lauryl sulfate, sodium doccusate,vitamin E TPGS, dimethylacetamide, N-methylpyrrolidone,N-hydroxyethylpyrrolidone, polyvinylpyrrolidone, hydroxypropylmethylcellulose, hydroxypropyl cyclodextrins, ethanol, n-butanol, isopropylalcohol, cholesterol, bile salts, polyethylene glycol 200-600,glycofurol, transcutol, propylene glycol, dimethyl isosorbide, and thelike.

Stabilizers include compounds such as any antioxidation agents, buffers,acids, preservatives, and the like.

Suspending agents include compounds such as polyvinylpyrrolidone (e.g.,polyvinylpyrrolidone K12, polyvinylpyrrolidone K17, polyvinylpyrrolidoneK25, or polyvinylpyrrolidone K30), vinyl pyrrolidone/vinyl acetatecopolymer (S630), polyethylene glycol (e.g., the polyethylene glycol hasa molecular weight of about 300 to about 6000, or about 3350 to about4000, or about 7000 to about 5400), sodium carboxymethylcellulose,methylcellulose, hydroxypropylmethylcellulose, hydroxymethylcelluloseacetate stearate, polysorbate-80, hydroxyethylcellulose, sodiumalginate, gums (such as, e.g., gum tragacanth and gum acacia, guar gum,xanthans, including xanthan gum), sugars, cellulosics (such as, e.g.,sodium carboxymethylcellulose, methylcellulose, sodiumcarboxymethylcellulose, hydroxypropylmethylcellulose,hydroxyethylcellulose), polysorbate-80, sodium alginate, polyethoxylatedsorbitan monolaurate, polyethoxylated sorbitan monolaurate, povidone,and the like.

Surfactants include compounds such as sodium lauryl sulfate, sodiumdocusate, Tween 60 or 80, triacetin, vitamin E TPGS, sorbitanmonooleate, polyoxyethylene sorbitan monooleate, polysorbates,polaxomers, bile salts, glyceryl monostearate, copolymers of ethyleneoxide and propylene oxide, e.g., Pluronic® (BASF), and the like.Additional surfactants include polyoxyethylene fatty acid glycerides andvegetable oils, e.g., polyoxyethylene (60) hydrogenated castor oil; andpolyoxyethylene alkylethers and alkylphenyl ethers, e.g., octoxynol 10,octoxynol 40. Sometimes, surfactants are included to enhance physicalstability or for other purposes.

Viscosity enhancing agents include, e.g., methyl cellulose, xanthan gum,carboxymethyl cellulose, hydroxypropyl cellulose, hydroxypropylmethylcellulose, hydroxypropylmethyl cellulose acetate stearate,hydroxypropylmethyl cellulose phthalate, carbomer, polyvinyl alcohol,alginates, acacia, chitosans, and combinations thereof.

Wetting agents include compounds such as oleic acid, glycerylmonostearate, sorbitan monooleate, sorbitan monolaurate, triethanolamineoleate, polyoxyethylene sorbitan monooleate, polyoxyethylene sorbitanmonolaurate, sodium docusate, sodium oleate, sodium lauryl sulfate,sodium doccusate, triacetin, Tween 80, vitamin E TPGS, ammonium salts,and the like.

Therapeutic Regimens

In some embodiments, the pharmaceutical compositions described hereinare administered for therapeutic applications. In some embodiments, thepharmaceutical composition is administered once per day, twice per day,three times per day, or more. The pharmaceutical composition isadministered daily, every day, every alternate day, five days a week,once a week, every other week, two weeks per month, three weeks permonth, once a month, twice a month, three times per month, or more. Thepharmaceutical composition is administered for at least 1 month, 2months, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9months, 10 months, 11 months, 12 months, 18 months, 2 years, 3 years, ormore.

In some embodiments, one or more pharmaceutical compositions areadministered simultaneously, sequentially, or at an interval period oftime. In some embodiments, one or more pharmaceutical compositions areadministered simultaneously. In some cases, one or more pharmaceuticalcompositions are administered sequentially. In additional cases, one ormore pharmaceutical compositions are administered at an interval periodof time (e.g., the first administration of a first pharmaceuticalcomposition is on day one followed by an interval of at least 1, 2, 3,4, 5, or more days prior to the administration of at least a secondpharmaceutical composition).

In some embodiments, two or more different pharmaceutical compositionsare coadministered. In some instances, the two or more differentpharmaceutical compositions are coadministered simultaneously. In somecases, the two or more different pharmaceutical compositions arecoadministered sequentially without a gap of time betweenadministrations. In other cases, the two or more differentpharmaceutical compositions are coadministered sequentially with a gapof about 0.5 hour, 1 hour, 2 hour, 3 hour, 12 hours, 1 day, 2 days, ormore between administrations.

In the case wherein the patient's status does improve, upon the doctor'sdiscretion the administration of the composition is given continuously;alternatively, the dose of the composition being administered istemporarily reduced or temporarily suspended for a certain length oftime (i.e., a “drug holiday”). In some instances, the length of the drugholiday varies between 2 days and 1 year, including by way of exampleonly, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 10 days, 12 days,15 days, 20 days, 28 days, 35 days, 50 days, 70 days, 100 days, 120days, 150 days, 180 days, 200 days, 250 days, 280 days, 300 days, 320days, 350 days, or 365 days. The dose reduction during a drug holiday isfrom 10%-100%, including, by way of example only, 10%, 15%, 20%, 25%,30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or100%.

Once improvement of the patient's conditions has occurred, a maintenancedose is administered if necessary. Subsequently, the dosage or thefrequency of administration, or both, are optionally reduced, as afunction of the symptoms, to a level at which the improved disease,disorder or condition is retained.

In some embodiments, the amount of a given agent that correspond to suchan amount varies depending upon factors such as the particular compound,the severity of the disease, the identity (e.g., weight) of the subjector host in need of treatment, but nevertheless is routinely determinedin a manner known in the art according to the particular circumstancessurrounding the case, including, e.g., the specific agent beingadministered, the route of administration, and the subject or host beingtreated. In some instances, the desired dose is conveniently presentedin a single dose or as divided doses administered simultaneously (orover a short period of time) or at appropriate intervals, for example astwo, three, four or more sub-doses per day.

The foregoing ranges are merely suggestive, as the number of variablesin regard to an individual treatment regime is large, and considerableexcursions from these recommended values are not uncommon. Such dosagesare altered depending on a number of variables, not limited to theactivity of the compound used, the disease or condition to be treated,the mode of administration, the requirements of the individual subject,the severity of the disease or condition being treated, and the judgmentof the practitioner.

In some embodiments, toxicity and therapeutic efficacy of suchtherapeutic regimens are determined by standard pharmaceuticalprocedures in cell cultures or experimental animals, including, but notlimited to, the determination of the LD50 (the dose lethal to 50% of thepopulation) and the ED50 (the dose therapeutically effective in 50% ofthe population). The dose ratio between the toxic and therapeuticeffects is the therapeutic index and it is expressed as the ratiobetween LD50 and ED50. Compounds exhibiting high therapeutic indices arepreferred. The data obtained from cell culture assays and animal studiesare used in formulating a range of dosage for use in humans. The dosageof such compounds lies preferably within a range of circulatingconcentrations that include the ED50 with minimal toxicity. The dosagevaries within this range depending upon the dosage form employed and theroute of administration utilized.

Kits/Article of Manufacture

Disclosed herein, in certain embodiments, are kits and articles ofmanufacture for use with one or more of the compositions and methodsdescribed herein. Such kits include a carrier, package, or containerthat is compartmentalized to receive one or more containers such asvials, tubes, and the like, each of the container(s) comprising one ofthe separate elements to be used in a method described herein. Suitablecontainers include, for example, bottles, vials, syringes, and testtubes. In one embodiment, the containers are formed from a variety ofmaterials such as glass or plastic.

The articles of manufacture provided herein contain packaging materials.Examples of pharmaceutical packaging materials include, but are notlimited to, blister packs, bottles, tubes, bags, containers, bottles,and any packaging material suitable for a selected formulation andintended mode of administration and treatment.

For example, the container(s) include EGFR nucleic acid moleculedescribed herein. Such kits optionally include an identifyingdescription or label or instructions relating to its use in the methodsdescribed herein.

A kit typically includes labels listing contents and/or instructions foruse and package inserts with instructions for use. A set of instructionswill also typically be included.

In one embodiment, a label is on or associated with the container. Inone embodiment, a label is on a container when letters, numbers, orother characters forming the label are attached, molded or etched intothe container itself; a label is associated with a container when it ispresent within a receptacle or carrier that also holds the container,e.g., as a package insert. In one embodiment, a label is used toindicate that the contents are to be used for a specific therapeuticapplication. The label also indicates directions for use of thecontents, such as in the methods described herein.

In certain embodiments, the pharmaceutical compositions are presented ina pack or dispenser device which contains one or more unit dosage formscontaining a compound provided herein. The pack, for example, containsmetal or plastic foil, such as a blister pack. In one embodiment, thepack or dispenser device is accompanied by instructions foradministration. In one embodiment, the pack or dispenser is alsoaccompanied with a notice associated with the container in a formprescribed by a governmental agency regulating the manufacture, use, orsale of pharmaceuticals, which notice is reflective of approval by theagency of the form of the drug for human or veterinary administration.Such notice, for example, is the labeling approved by the U.S. Food andDrug Administration for prescription drugs, or the approved productinsert. In one embodiment, compositions containing a compound providedherein formulated in a compatible pharmaceutical carrier are alsoprepared, placed in an appropriate container, and labeled for treatmentof an indicated condition.

Certain Terminology

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as is commonly understood by one of skill in theart to which the claimed subject matter belongs. It is to be understoodthat the general description and the detailed description are exemplaryand explanatory only and are not restrictive of any subject matterclaimed. In this application, the use of the singular includes theplural unless specifically stated otherwise. It must be noted that, asused in the specification, the singular forms “a,” “an” and “the”include plural referents unless the context clearly dictates otherwise.In this application, the use of “or” means “and/or” unless statedotherwise. Furthermore, use of the term “including” as well as otherforms, such as “include”, “includes,” and “included,” is not limiting.

As used herein, ranges and amounts can be expressed as “about” aparticular value or range. About also includes the exact amount. Hence“about 5 μL” means “about 5 μL” and also “5 μL.” Generally, the term“about” includes an amount that is expected to be within experimentalerror, e.g., ±5%, ±10%, or ±15%.

The section headings used herein are for organizational purposes onlyand are not to be construed as limiting the subject matter described.

As used herein, the terms “individual(s)”, “subject(s)” and “patient(s)”mean any mammal. In some embodiments, the mammal is a human. In someembodiments, the mammal is a non-human. None of the terms require or arelimited to situations characterized by the supervision (e.g. constant orintermittent) of a health care worker (e.g. a doctor, a registerednurse, a nurse practitioner, a physician's assistant, an orderly or ahospice worker).

EXAMPLES

These examples are provided for illustrative purposes only and not tolimit the scope of the claims provided herein.

Example 1. Sequences

Table 1 illustrates target sequences described herein. Tables 2, 3, and6 illustrate polynucleic acid molecule sequences described herein.

TABLE 1 EGFR Target Sequences 19mer pos. SEQ  in sequence of total 23merID hs Id # NM_005228.3 target site in NM_005228.3 NO: 68 68-86GGCGGCCGGAGUCCCGAGCUAGC 1 71 71-89 GGCCGGAGUCCCGAGCUAGCCCC 2 72 72-90GCCGGAGUCCCGAGCUAGCCCCG 3 73 73-91 CCGGAGUCCCGAGCUAGCCCCGG 4 74 74-92CGGAGUCCCGAGCUAGCCCCGGC 5 75 75-93 GGAGUCCCGAGCUAGCCCCGGCG 6 76 76-94GAGUCCCGAGCUAGCCCCGGCGG 7 78 78-96 GUCCCGAGCUAGCCCCGGCGGCC 8 114 114-132CCGGACGACAGGCCACCUCGUCG 9 115 115-133 CGGACGACAGGCCACCUCGUCGG 10 116116-134 GGACGACAGGCCACCUCGUCGGC 11 117 117-135 GACGACAGGCCACCUCGUCGGCG12 118 118-136 ACGACAGGCCACCUCGUCGGCGU 13 120 120-138GACAGGCCACCUCGUCGGCGUCC 14 121 121-139 ACAGGCCACCUCGUCGGCGUCCG 15 122122-140 CAGGCCACCUCGUCGGCGUCCGC 16 123 123-141 AGGCCACCUCGUCGGCGUCCGCC17 124 124-142 GGCCACCUCGUCGGCGUCCGCCC 18 125 125-143GCCACCUCGUCGGCGUCCGCCCG 19 126 126-144 CCACCUCGUCGGCGUCCGCCCGA 20 127127-145 CACCUCGUCGGCGUCCGCCCGAG 21 128 128-146 ACCUCGUCGGCGUCCGCCCGAGU22 129 129-147 CCUCGUCGGCGUCCGCCCGAGUC 23 130 130-148CUCGUCGGCGUCCGCCCGAGUCC 24 131 131-149 UCGUCGGCGUCCGCCCGAGUCCC 25 132132-150 CGUCGGCGUCCGCCCGAGUCCCC 26 135 135-153 CGGCGUCCGCCCGAGUCCCCGCC27 136 136-154 GGCGUCCGCCCGAGUCCCCGCCU 28 141 141-159CCGCCCGAGUCCCCGCCUCGCCG 29 164 164-182 CCAACGCCACAACCACCGCGCAC 30 165165-183 CAACGCCACAACCACCGCGCACG 31 166 166-184 AACGCCACAACCACCGCGCACGG32 168 168-186 CGCCACAACCACCGCGCACGGCC 33 169 169-187GCCACAACCACCGCGCACGGCCC 34 170 170-188 CCACAACCACCGCGCACGGCCCC 35 247247-265 CGAUGCGACCCUCCGGGACGGCC 36 248 248-266 GAUGCGACCCUCCGGGACGGCCG37 249 249-267 AUGCGACCCUCCGGGACGGCCGG 38 251 251-269GCGACCCUCCGGGACGGCCGGGG 39 252 252-270 CGACCCUCCGGGACGGCCGGGGC 40 254254-272 ACCCUCCGGGACGGCCGGGGCAG 41 329 329-347 AAAGAAAGUUUGCCAAGGCACGA42 330 330-348 AAGAAAGUUUGCCAAGGCACGAG 43 332 332-350GAAAGUUUGCCAAGGCACGAGUA 44 333 333-351 AAAGUUUGCCAAGGCACGAGUAA 45 334334-352 AAGUUUGCCAAGGCACGAGUAAC 46 335 335-353 AGUUUGCCAAGGCACGAGUAACA47 336 336-354 GUUUGCCAAGGCACGAGUAACAA 48 337 337-355UUUGCCAAGGCACGAGUAACAAG 49 338 338-356 UUGCCAAGGCACGAGUAACAAGC 50 361361-379 UCACGCAGUUGGGCACUUUUGAA 51 362 362-380 CACGCAGUUGGGCACUUUUGAAG52 363 363-381 ACGCAGUUGGGCACUUUUGAAGA 53 364 364-382CGCAGUUGGGCACUUUUGAAGAU 54 365 365-383 GCAGUUGGGCACUUUUGAAGAUC 55 366366-384 CAGUUGGGCACUUUUGAAGAUCA 56 367 367-385 AGUUGGGCACUUUUGAAGAUCAU57 368 368-386 GUUGGGCACUUUUGAAGAUCAUU 58 369 369-387UUGGGCACUUUUGAAGAUCAUUU 59 377 377-395 UUUUGAAGAUCAUUUUCUCAGCC 60 379379-397 UUGAAGAUCAUUUUCUCAGCCUC 61 380 380-398 UGAAGAUCAUUUUCUCAGCCUCC62 385 385-403 AUCAUUUUCUCAGCCUCCAGAGG 63 394 394-412UCAGCCUCCAGAGGAUGUUCAAU 64 396 396-414 AGCCUCCAGAGGAUGUUCAAUAA 65 397397-415 GCCUCCAGAGGAUGUUCAAUAAC 66 401 401-419 CCAGAGGAUGUUCAAUAACUGUG67 403 403-421 AGAGGAUGUUCAAUAACUGUGAG 68 407 407-425GAUGUUCAAUAACUGUGAGGUGG 69 409 409-427 UGUUCAAUAACUGUGAGGUGGUC 70 410410-428 GUUCAAUAACUGUGAGGUGGUCC 71 411 411-429 UUCAAUAACUGUGAGGUGGUCCU72 412 412-430 UCAAUAACUGUGAGGUGGUCCUU 73 413 413-431CAAUAACUGUGAGGUGGUCCUUG 74 414 414-432 AAUAACUGUGAGGUGGUCCUUGG 75 416416-434 UAACUGUGAGGUGGUCCUUGGGA 76 418 418-436 ACUGUGAGGUGGUCCUUGGGAAU77 419 419-437 CUGUGAGGUGGUCCUUGGGAAUU 78 425 425-443GGUGGUCCUUGGGAAUUUGGAAA 79 431 431-449 CCUUGGGAAUUUGGAAAUUACCU 80 432432-450 CUUGGGAAUUUGGAAAUUACCUA 81 433 433-451 UUGGGAAUUUGGAAAUUACCUAU82 434 434-452 UGGGAAUUUGGAAAUUACCUAUG 83 458 458-476GCAGAGGAAUUAUGAUCUUUCCU 84 459 459-477 CAGAGGAAUUAUGAUCUUUCCUU 85 463463-481 GGAAUUAUGAUCUUUCCUUCUUA 86 464 464-482 GAAUUAUGAUCUUUCCUUCUUAA87 466 466-484 AUUAUGAUCUUUCCUUCUUAAAG 88 468 468-486UAUGAUCUUUCCUUCUUAAAGAC 89 471 471-489 GAUCUUUCCUUCUUAAAGACCAU 90 476476-494 UUCCUUCUUAAAGACCAUCCAGG 91 477 477-495 UCCUUCUUAAAGACCAUCCAGGA92 479 479-497 CUUCUUAAAGACCAUCCAGGAGG 93 481 481-499UCUUAAAGACCAUCCAGGAGGUG 94 482 482-500 CUUAAAGACCAUCCAGGAGGUGG 95 492492-510 AUCCAGGAGGUGGCUGGUUAUGU 96 493 493-511 UCCAGGAGGUGGCUGGUUAUGUC97 494 494-512 CCAGGAGGUGGCUGGUUAUGUCC 98 495 495-513CAGGAGGUGGCUGGUUAUGUCCU 99 496 496-514 AGGAGGUGGCUGGUUAUGUCCUC 100 497497-515 GGAGGUGGCUGGUUAUGUCCUCA 101 499 499-517 AGGUGGCUGGUUAUGUCCUCAUU102 520 520-538 UUGCCCUCAACACAGUGGAGCGA 103 542 542-560AAUUCCUUUGGAAAACCUGCAGA 104 543 543-561 AUUCCUUUGGAAAACCUGCAGAU 105 550550-568 UGGAAAACCUGCAGAUCAUCAGA 106 551 551-569 GGAAAACCUGCAGAUCAUCAGAG107 553 553-571 AAAACCUGCAGAUCAUCAGAGGA 108 556 556-574ACCUGCAGAUCAUCAGAGGAAAU 109 586 586-604 ACGAAAAUUCCUAUGCCUUAGCA 110 587587-605 CGAAAAUUCCUAUGCCUUAGCAG 111 589 589-607 AAAAUUCCUAUGCCUUAGCAGUC112 592 592-610 AUUCCUAUGCCUUAGCAGUCUUA 113 593 593-611UUCCUAUGCCUUAGCAGUCUUAU 114 594 594-612 UCCUAUGCCUUAGCAGUCUUAUC 115 596596-614 CUAUGCCUUAGCAGUCUUAUCUA 116 597 597-615 UAUGCCUUAGCAGUCUUAUCUAA117 598 598-616 AUGCCUUAGCAGUCUUAUCUAAC 118 599 599-617UGCCUUAGCAGUCUUAUCUAACU 119 600 600-618 GCCUUAGCAGUCUUAUCUAACUA 120 601601-619 CCUUAGCAGUCUUAUCUAACUAU 121 602 602-620 CUUAGCAGUCUUAUCUAACUAUG122 603 603-621 UUAGCAGUCUUAUCUAACUAUGA 123 604 604-622UAGCAGUCUUAUCUAACUAUGAU 124 605 605-623 AGCAGUCUUAUCUAACUAUGAUG 125 608608-626 AGUCUUAUCUAACUAUGAUGCAA 126 609 609-627 GUCUUAUCUAACUAUGAUGCAAA127 610 610-628 UCUUAUCUAACUAUGAUGCAAAU 128 611 611-629CUUAUCUAACUAUGAUGCAAAUA 129 612 612-630 UUAUCUAACUAUGAUGCAAAUAA 130 613613-631 UAUCUAACUAUGAUGCAAAUAAA 131 614 614-632 AUCUAACUAUGAUGCAAAUAAAA132 616 616-634 CUAACUAUGAUGCAAAUAAAACC 133 622 622-640AUGAUGCAAAUAAAACCGGACUG 134 623 623-641 UGAUGCAAAUAAAACCGGACUGA 135 624624-642 GAUGCAAAUAAAACCGGACUGAA 136 626 626-644 UGCAAAUAAAACCGGACUGAAGG137 627 627-645 GCAAAUAAAACCGGACUGAAGGA 138 628 628-646CAAAUAAAACCGGACUGAAGGAG 139 630 630-648 AAUAAAACCGGACUGAAGGAGCU 140 631631-649 AUAAAACCGGACUGAAGGAGCUG 141 632 632-650 UAAAACCGGACUGAAGGAGCUGC142 633 633-651 AAAACCGGACUGAAGGAGCUGCC 143 644 644-662GAAGGAGCUGCCCAUGAGAAAUU 144 665 665-683 UUUACAGGAAAUCCUGCAUGGCG 145 668668-686 ACAGGAAAUCCUGCAUGGCGCCG 146 669 669-687 CAGGAAAUCCUGCAUGGCGCCGU147 670 670-688 AGGAAAUCCUGCAUGGCGCCGUG 148 671 671-689GGAAAUCCUGCAUGGCGCCGUGC 149 672 672-690 GAAAUCCUGCAUGGCGCCGUGCG 150 674674-692 AAUCCUGCAUGGCGCCGUGCGGU 151 676 676-694 UCCUGCAUGGCGCCGUGCGGUUC152 677 677-695 CCUGCAUGGCGCCGUGCGGUUCA 153 678 678-696CUGCAUGGCGCCGUGCGGUUCAG 154 680 680-698 GCAUGGCGCCGUGCGGUUCAGCA 155 681681-699 CAUGGCGCCGUGCGGUUCAGCAA 156 682 682-700 AUGGCGCCGUGCGGUUCAGCAAC157 683 683-701 UGGCGCCGUGCGGUUCAGCAACA 158 684 684-702GGCGCCGUGCGGUUCAGCAACAA 159 685 685-703 GCGCCGUGCGGUUCAGCAACAAC 160 686686-704 CGCCGUGCGGUUCAGCAACAACC 161 688 688-706 CCGUGCGGUUCAGCAACAACCCU162 690 690-708 GUGCGGUUCAGCAACAACCCUGC 163 692 692-710GCGGUUCAGCAACAACCCUGCCC 164 698 698-716 CAGCAACAACCCUGCCCUGUGCA 165 700700-718 GCAACAACCCUGCCCUGUGCAAC 166 719 719-737 CAACGUGGAGAGCAUCCAGUGGC167 720 720-738 AACGUGGAGAGCAUCCAGUGGCG 168 721 721-739ACGUGGAGAGCAUCCAGUGGCGG 169 724 724-742 UGGAGAGCAUCCAGUGGCGGGAC 170 725725-743 GGAGAGCAUCCAGUGGCGGGACA 171 726 726-744 GAGAGCAUCCAGUGGCGGGACAU172 733 733-751 UCCAGUGGCGGGACAUAGUCAGC 173 734 734-752CCAGUGGCGGGACAUAGUCAGCA 174 736 736-754 AGUGGCGGGACAUAGUCAGCAGU 175 737737-755 GUGGCGGGACAUAGUCAGCAGUG 176 763 763-781 UUCUCAGCAACAUGUCGAUGGAC177 765 765-783 CUCAGCAACAUGUCGAUGGACUU 178 766 766-784UCAGCAACAUGUCGAUGGACUUC 179 767 767-785 CAGCAACAUGUCGAUGGACUUCC 180 769769-787 GCAACAUGUCGAUGGACUUCCAG 181 770 770-788 CAACAUGUCGAUGGACUUCCAGA182 771 771-789 AACAUGUCGAUGGACUUCCAGAA 183 772 772-790ACAUGUCGAUGGACUUCCAGAAC 184 775 775-793 UGUCGAUGGACUUCCAGAACCAC 185 789789-807 CAGAACCACCUGGGCAGCUGCCA 186 798 798-816 CUGGGCAGCUGCCAAAAGUGUGA187 800 800-818 GGGCAGCUGCCAAAAGUGUGAUC 188 805 805-823GCUGCCAAAAGUGUGAUCCAAGC 189 806 806-824 CUGCCAAAAGUGUGAUCCAAGCU 190 807807-825 UGCCAAAAGUGUGAUCCAAGCUG 191 810 810-828 CAAAAGUGUGAUCCAAGCUGUCC192 814 814-832 AGUGUGAUCCAAGCUGUCCCAAU 193 815 815-833GUGUGAUCCAAGCUGUCCCAAUG 194 817 817-835 GUGAUCCAAGCUGUCCCAAUGGG 195 818818-836 UGAUCCAAGCUGUCCCAAUGGGA 196 819 819-837 GAUCCAAGCUGUCCCAAUGGGAG197 820 820-838 AUCCAAGCUGUCCCAAUGGGAGC 198 821 821-839UCCAAGCUGUCCCAAUGGGAGCU 199 823 823-841 CAAGCUGUCCCAAUGGGAGCUGC 200 826826-844 GCUGUCCCAAUGGGAGCUGCUGG 201 847 847-865 GGGGUGCAGGAGAGGAGAACUGC202 871 871-889 AGAAACUGACCAAAAUCAUCUGU 203 872 872-890GAAACUGACCAAAAUCAUCUGUG 204 873 873-891 AAACUGACCAAAAUCAUCUGUGC 205 877877-895 UGACCAAAAUCAUCUGUGCCCAG 206 878 878-896 GACCAAAAUCAUCUGUGCCCAGC207 881 881-899 CAAAAUCAUCUGUGCCCAGCAGU 208 890 890-908CUGUGCCCAGCAGUGCUCCGGGC 209 892 892-910 GUGCCCAGCAGUGCUCCGGGCGC 210 929929-947 CCCCAGUGACUGCUGCCACAACC 211 930 930-948 CCCAGUGACUGCUGCCACAACCA212 979 979-997 GGGAGAGCGACUGCCUGGUCUGC 213 980 980-998GGAGAGCGACUGCCUGGUCUGCC 214 981 981-999 GAGAGCGACUGCCUGGUCUGCCG 215 982 982-1000 AGAGCGACUGCCUGGUCUGCCGC 216 983  983-1001GAGCGACUGCCUGGUCUGCCGCA 217 984  984-1002 AGCGACUGCCUGGUCUGCCGCAA 218989  989-1007 CUGCCUGGUCUGCCGCAAAUUCC 219 990  990-1008UGCCUGGUCUGCCGCAAAUUCCG 220 991  991-1009 GCCUGGUCUGCCGCAAAUUCCGA 221992  992-1010 CCUGGUCUGCCGCAAAUUCCGAG 222 994  994-1012UGGUCUGCCGCAAAUUCCGAGAC 223 995  995-1013 GGUCUGCCGCAAAUUCCGAGACG 224996  996-1014 GUCUGCCGCAAAUUCCGAGACGA 225 997  997-1015UCUGCCGCAAAUUCCGAGACGAA 226 999  999-1017 UGCCGCAAAUUCCGAGACGAAGC 2271004 1004-1022 CAAAUUCCGAGACGAAGCCACGU 228 1005 1005-1023AAAUUCCGAGACGAAGCCACGUG 229 1006 1006-1024 AAUUCCGAGACGAAGCCACGUGC 2301007 1007-1025 AUUCCGAGACGAAGCCACGUGCA 231 1008 1008-1026UUCCGAGACGAAGCCACGUGCAA 232 1010 1010-1028 CCGAGACGAAGCCACGUGCAAGG 2331013 1013-1031 AGACGAAGCCACGUGCAAGGACA 234 1014 1014-1032GACGAAGCCACGUGCAAGGACAC 235 1015 1015-1033 ACGAAGCCACGUGCAAGGACACC 2361016 1016-1034 CGAAGCCACGUGCAAGGACACCU 237 1040 1040-1058CCCCCCACUCAUGCUCUACAACC 238 1042 1042-1060 CCCCACUCAUGCUCUACAACCCC 2391044 1044-1062 CCACUCAUGCUCUACAACCCCAC 240 1047 1047-1065CUCAUGCUCUACAACCCCACCAC 241 1071 1071-1089 UACCAGAUGGAUGUGAACCCCGA 2421073 1073-1091 CCAGAUGGAUGUGAACCCCGAGG 243 1074 1074-1092CAGAUGGAUGUGAACCCCGAGGG 244 1075 1075-1093 AGAUGGAUGUGAACCCCGAGGGC 2451077 1077-1095 AUGGAUGUGAACCCCGAGGGCAA 246 1078 1078-1096UGGAUGUGAACCCCGAGGGCAAA 247 1080 1080-1098 GAUGUGAACCCCGAGGGCAAAUA 2481084 1084-1102 UGAACCCCGAGGGCAAAUACAGC 249 1085 1085-1103GAACCCCGAGGGCAAAUACAGCU 250 1087 1087-1105 ACCCCGAGGGCAAAUACAGCUUU 2511088 1088-1106 CCCCGAGGGCAAAUACAGCUUUG 252 1089 1089-1107CCCGAGGGCAAAUACAGCUUUGG 253 1096 1096-1114 GCAAAUACAGCUUUGGUGCCACC 2541097 1097-1115 CAAAUACAGCUUUGGUGCCACCU 255 1098 1098-1116AAAUACAGCUUUGGUGCCACCUG 256 1104 1104-1122 AGCUUUGGUGCCACCUGCGUGAA 2571106 1106-1124 CUUUGGUGCCACCUGCGUGAAGA 258 1112 1112-1130UGCCACCUGCGUGAAGAAGUGUC 259 1116 1116-1134 ACCUGCGUGAAGAAGUGUCCCCG 2601117 1117-1135 CCUGCGUGAAGAAGUGUCCCCGU 261 1118 1118-1136CUGCGUGAAGAAGUGUCCCCGUA 262 1119 1119-1137 UGCGUGAAGAAGUGUCCCCGUAA 2631120 1120-1138 GCGUGAAGAAGUGUCCCCGUAAU 264 1121 1121-1139CGUGAAGAAGUGUCCCCGUAAUU 265 1122 1122-1140 GUGAAGAAGUGUCCCCGUAAUUA 2661123 1123-1141 UGAAGAAGUGUCCCCGUAAUUAU 267 1124 1124-1142GAAGAAGUGUCCCCGUAAUUAUG 268 1125 1125-1143 AAGAAGUGUCCCCGUAAUUAUGU 2691126 1126-1144 AGAAGUGUCCCCGUAAUUAUGUG 270 1127 1127-1145GAAGUGUCCCCGUAAUUAUGUGG 271 1128 1128-1146 AAGUGUCCCCGUAAUUAUGUGGU 2721129 1129-1147 AGUGUCCCCGUAAUUAUGUGGUG 273 1130 1130-1148GUGUCCCCGUAAUUAUGUGGUGA 274 1132 1132-1150 GUCCCCGUAAUUAUGUGGUGACA 2751134 1134-1152 CCCCGUAAUUAUGUGGUGACAGA 276 1136 1136-1154CCGUAAUUAUGUGGUGACAGAUC 277 1137 1137-1155 CGUAAUUAUGUGGUGACAGAUCA 2781138 1138-1156 GUAAUUAUGUGGUGACAGAUCAC 279 1139 1139-1157UAAUUAUGUGGUGACAGAUCACG 280 1140 1140-1158 AAUUAUGUGGUGACAGAUCACGG 2811142 1142-1160 UUAUGUGGUGACAGAUCACGGCU 282 1145 1145-1163UGUGGUGACAGAUCACGGCUCGU 283 1147 1147-1165 UGGUGACAGAUCACGGCUCGUGC 2841148 1148-1166 GGUGACAGAUCACGGCUCGUGCG 285 1149 1149-1167GUGACAGAUCACGGCUCGUGCGU 286 1150 1150-1168 UGACAGAUCACGGCUCGUGCGUC 2871151 1151-1169 GACAGAUCACGGCUCGUGCGUCC 288 1152 1152-1170ACAGAUCACGGCUCGUGCGUCCG 289 1153 1153-1171 CAGAUCACGGCUCGUGCGUCCGA 2901154 1154-1172 AGAUCACGGCUCGUGCGUCCGAG 291 1155 1155-1173GAUCACGGCUCGUGCGUCCGAGC 292 1156 1156-1174 AUCACGGCUCGUGCGUCCGAGCC 2931157 1157-1175 UCACGGCUCGUGCGUCCGAGCCU 294 1160 1160-1178CGGCUCGUGCGUCCGAGCCUGUG 295 1200 1200-1218 AUGGAGGAAGACGGCGUCCGCAA 2961201 1201-1219 UGGAGGAAGACGGCGUCCGCAAG 297 1203 1203-1221GAGGAAGACGGCGUCCGCAAGUG 298 1204 1204-1222 AGGAAGACGGCGUCCGCAAGUGU 2991205 1205-1223 GGAAGACGGCGUCCGCAAGUGUA 300 1207 1207-1225AAGACGGCGUCCGCAAGUGUAAG 301 1208 1208-1226 AGACGGCGUCCGCAAGUGUAAGA 3021211 1211-1229 CGGCGUCCGCAAGUGUAAGAAGU 303 1212 1212-1230GGCGUCCGCAAGUGUAAGAAGUG 304 1213 1213-1231 GCGUCCGCAAGUGUAAGAAGUGC 3051214 1214-1232 CGUCCGCAAGUGUAAGAAGUGCG 306 1215 1215-1233GUCCGCAAGUGUAAGAAGUGCGA 307 1216 1216-1234 UCCGCAAGUGUAAGAAGUGCGAA 3081217 1217-1235 CCGCAAGUGUAAGAAGUGCGAAG 309 1219 1219-1237GCAAGUGUAAGAAGUGCGAAGGG 310 1220 1220-1238 CAAGUGUAAGAAGUGCGAAGGGC 3111221 1221-1239 AAGUGUAAGAAGUGCGAAGGGCC 312 1222 1222-1240AGUGUAAGAAGUGCGAAGGGCCU 313 1223 1223-1241 GUGUAAGAAGUGCGAAGGGCCUU 3141224 1224-1242 UGUAAGAAGUGCGAAGGGCCUUG 315 1225 1225-1243GUAAGAAGUGCGAAGGGCCUUGC 316 1226 1226-1244 UAAGAAGUGCGAAGGGCCUUGCC 3171229 1229-1247 GAAGUGCGAAGGGCCUUGCCGCA 318 1230 1230-1248AAGUGCGAAGGGCCUUGCCGCAA 319 1231 1231-1249 AGUGCGAAGGGCCUUGCCGCAAA 3201232 1232-1250 GUGCGAAGGGCCUUGCCGCAAAG 321 1233 1233-1251UGCGAAGGGCCUUGCCGCAAAGU 322 1235 1235-1253 CGAAGGGCCUUGCCGCAAAGUGU 3231236 1236-1254 GAAGGGCCUUGCCGCAAAGUGUG 324 1237 1237-1255AAGGGCCUUGCCGCAAAGUGUGU 325 1238 1238-1256 AGGGCCUUGCCGCAAAGUGUGUA 3261239 1239-1257 GGGCCUUGCCGCAAAGUGUGUAA 327 1241 1241-1259GCCUUGCCGCAAAGUGUGUAACG 328 1261 1261-1279 ACGGAAUAGGUAUUGGUGAAUUU 3291262 1262-1280 CGGAAUAGGUAUUGGUGAAUUUA 330 1263 1263-1281GGAAUAGGUAUUGGUGAAUUUAA 331 1264 1264-1282 GAAUAGGUAUUGGUGAAUUUAAA 3321266 1266-1284 AUAGGUAUUGGUGAAUUUAAAGA 333 1267 1267-1285UAGGUAUUGGUGAAUUUAAAGAC 334 1289 1289-1307 CUCACUCUCCAUAAAUGCUACGA 3351313 1313-1331 UAUUAAACACUUCAAAAACUGCA 336 1320 1320-1338CACUUCAAAAACUGCACCUCCAU 337 1321 1321-1339 ACUUCAAAAACUGCACCUCCAUC 3381322 1322-1340 CUUCAAAAACUGCACCUCCAUCA 339 1323 1323-1341UUCAAAAACUGCACCUCCAUCAG 340 1324 1324-1342 UCAAAAACUGCACCUCCAUCAGU 3411328 1328-1346 AAACUGCACCUCCAUCAGUGGCG 342 1332 1332-1350UGCACCUCCAUCAGUGGCGAUCU 343 1333 1333-1351 GCACCUCCAUCAGUGGCGAUCUC 3441335 1335-1353 ACCUCCAUCAGUGGCGAUCUCCA 345 1338 1338-1356UCCAUCAGUGGCGAUCUCCACAU 346 1344 1344-1362 AGUGGCGAUCUCCACAUCCUGCC 3471345 1345-1363 GUGGCGAUCUCCACAUCCUGCCG 348 1346 1346-1364UGGCGAUCUCCACAUCCUGCCGG 349 1347 1347-1365 GGCGAUCUCCACAUCCUGCCGGU 3501348 1348-1366 GCGAUCUCCACAUCCUGCCGGUG 351 1353 1353-1371CUCCACAUCCUGCCGGUGGCAUU 352 1354 1354-1372 UCCACAUCCUGCCGGUGGCAUUU 3531355 1355-1373 CCACAUCCUGCCGGUGGCAUUUA 354 1357 1357-1375ACAUCCUGCCGGUGGCAUUUAGG 355 1360 1360-1378 UCCUGCCGGUGGCAUUUAGGGGU 3561361 1361-1379 CCUGCCGGUGGCAUUUAGGGGUG 357 1362 1362-1380CUGCCGGUGGCAUUUAGGGGUGA 358 1363 1363-1381 UGCCGGUGGCAUUUAGGGGUGAC 3591366 1366-1384 CGGUGGCAUUUAGGGGUGACUCC 360 1369 1369-1387UGGCAUUUAGGGGUGACUCCUUC 361 1370 1370-1388 GGCAUUUAGGGGUGACUCCUUCA 3621371 1371-1389 GCAUUUAGGGGUGACUCCUUCAC 363 1372 1372-1390CAUUUAGGGGUGACUCCUUCACA 364 1373 1373-1391 AUUUAGGGGUGACUCCUUCACAC 3651374 1374-1392 UUUAGGGGUGACUCCUUCACACA 366 1404 1404-1422CCUCUGGAUCCACAGGAACUGGA 367 1408 1408-1426 UGGAUCCACAGGAACUGGAUAUU 3681409 1409-1427 GGAUCCACAGGAACUGGAUAUUC 369 1411 1411-1429AUCCACAGGAACUGGAUAUUCUG 370 1412 1412-1430 UCCACAGGAACUGGAUAUUCUGA 3711419 1419-1437 GAACUGGAUAUUCUGAAAACCGU 372 1426 1426-1444AUAUUCUGAAAACCGUAAAGGAA 373 1427 1427-1445 UAUUCUGAAAACCGUAAAGGAAA 3741430 1430-1448 UCUGAAAACCGUAAAGGAAAUCA 375 1431 1431-1449CUGAAAACCGUAAAGGAAAUCAC 376

TABLE 2 EGFR siRNA Sequences Sequence position SEQ SEQ  insense strand sequence  ID antisense strand sequence ID hs Id #NM_005228.3 (5′-3′) NO: (5′-3′) NO: 68 68-86 CGGCCGGAGUCCCGAG 377UAGCUCGGGACUCCGGC 378 CUATT CGTT 71 71-89 CCGGAGUCCCGAGCUA 379GGCUAGCUCGGGACUCC 380 GCCTT GGTT 72 72-90 CGGAGUCCCGAGCUAG 381GGGCUAGCUCGGGACUC 382 CCCTT CGTT 73 73-91 GGAGUCCCGAGCUAGC 383GGGGCUAGCUCGGGACU 384 CCCTT CCTT 74 74-92 GAGUCCCGAGCUAGCC 385CGGGGCUAGCUCGGGAC 386 CCGTT UCTT 75 75-93 AGUCCCGAGCUAGCCC 387CCGGGGCUAGCUCGGGA 388 CGGTT CUTT 76 76-94 GUCCCGAGCUAGCCCC 389GCCGGGGCUAGCUCGGG 390 GGCTT ACTT 78 78-96 CCCGAGCUAGCCCCGG 391CCGCCGGGGCUAGCUCG 392 CGGTT GGTT 114 114-132 GGACGACAGGCCACCU 393ACGAGGUGGCCUGUCGU 394 CGUTT CCTT 115 115-133 GACGACAGGCCACCUC 395GACGAGGUGGCCUGUCG 396 GUCTT UCTT 116 116-134 ACGACAGGCCACCUCG 397CGACGAGGUGGCCUGUC 398 UCGTT GUTT 117 117-135 CGACAGGCCACCUCGU 399CCGACGAGGUGGCCUGU 400 CGGTT CGTT 118 118-136 GACAGGCCACCUCGUC 401GCCGACGAGGUGGCCUG 402 GGCTT UCTT 120 120-138 CAGGCCACCUCGUCGG 403ACGCCGACGAGGUGGCC 404 CGUTT UGTT 121 121-139 AGGCCACCUCGUCGGC 405GACGCCGACGAGGUGGC 406 GUCTT CUTT 122 122-140 GGCCACCUCGUCGGCG 407GGACGCCGACGAGGUGG 408 UCCTT CCTT 123 123-141 GCCACCUCGUCGGCGU 409CGGACGCCGACGAGGUG 410 CCGTT GCTT 124 124-142 CCACCUCGUCGGCGUC 411GCGGACGCCGACGAGGU 412 CGCTT GGTT 125 125-143 CACCUCGUCGGCGUCC 413GGCGGACGCCGACGAGG 414 GCCTT UGTT 126 126-144 ACCUCGUCGGCGUCCG 415GGGCGGACGCCGACGAG 416 CCCTT GUTT 127 127-145 CCUCGUCGGCGUCCGC 417CGGGCGGACGCCGACGA 418 CCGTT GGTT 128 128-146 CUCGUCGGCGUCCGCC 419UCGGGCGGACGCCGACG 420 CGATT AGTT 129 129-147 UCGUCGGCGUCCGCCC 421CUCGGGCGGACGCCGAC 422 GAGTT GATT 130 130-148 CGUCGGCGUCCGCCCG 423ACUCGGGCGGACGCCGA 424 AGUTT CGTT 131 131-149 GUCGGCGUCCGCCCGA 425GACUCGGGCGGACGCCG 426 GUCTT ACTT 132 132-150 UCGGCGUCCGCCCGAG 427GGACUCGGGCGGACGCC 428 UCCTT GATT 135 135-153 GCGUCCGCCCGAGUCC 429CGGGGACUCGGGCGGAC 430 CCGTT GCTT 136 136-154 CGUCCGCCCGAGUCCC 431GCGGGGACUCGGGCGGA 432 CGCTT CGTT 141 141-159 GCCCGAGUCCCCGCCU 433GCGAGGCGGGGACUCGG 434 CGCTT GCTT 164 164-182 AACGCCACAACCACCG 435GCGCGGUGGUUGUGGC 436 CGCTT GUUTT 165 165-183 ACGCCACAACCACCGC 437UGCGCGGUGGUUGUGG 438 GCATT CGUTT 166 166-184 CGCCACAACCACCGCG 439GUGCGCGGUGGUUGUG 440 CACTT GCGTT 168 168-186 CCACAACCACCGCGCA 441CCGUGCGCGGUGGUUGU 442 CGGTT GGTT 169 169-187 CACAACCACCGCGCAC 443GCCGUGCGCGGUGGUUG 444 GGCTT UGTT 170 170-188 ACAACCACCGCGCACG 445GGCCGUGCGCGGUGGUU 446 GCCTT GUTT 247 247-265 AUGCGACCCUCCGGGA 447CCGUCCCGGAGGGUCGC 448 CGGTT AUTT 248 248-266 UGCGACCCUCCGGGAC 449GCCGUCCCGGAGGGUCG 450 GGCTT CATT 249 249-267 GCGACCCUCCGGGACG 451GGCCGUCCCGGAGGGUC 452 GCCTT GCTT 251 251-269 GACCCUCCGGGACGGC 453CCGGCCGUCCCGGAGGG 454 CGGTT UCTT 252 252-270 ACCCUCCGGGACGGCC 455CCCGGCCGUCCCGGAGG 456 GGGTT GUTT 254 254-272 CCUCCGGGACGGCCGG 457GCCCCGGCCGUCCCGGA 458 GGCTT GGTT 329 329-347 AGAAAGUUUGCCAAG 459GUGCCUUGGCAAACUUU 460 GCACTT CUTT 330 330-348 GAAAGUUUGCCAAGG 461CGUGCCUUGGCAAACUU 462 CACGTT UCTT 332 332-350 AAGUUUGCCAAGGCA 463CUCGUGCCUUGGCAAAC 464 CGAGTT UUTT 333 333-351 AGUUUGCCAAGGCAC 465ACUCGUGCCUUGGCAAA 466 GAGUTT CUTT 334 334-352 GUUUGCCAAGGCACG 467UACUCGUGCCUUGGCAA 468 AGUATT ACTT 335 335-353 UUUGCCAAGGCACGA 469UUACUCGUGCCUUGGCA 470 GUAATT AATT 336 336-354 UUGCCAAGGCACGAG 471GUUACUCGUGCCUUGGC 472 UAACTT AATT 337 337-355 UGCCAAGGCACGAGU 473UGUUACUCGUGCCUUGG 474 AACATT CATT 338 338-356 GCCAAGGCACGAGUA 475UUGUUACUCGUGCCUUG 476 ACAATT GCTT 361 361-379 ACGCAGUUGGGCACU 477CAAAAGUGCCCAACUGC 478 UUUGTT GUTT 362 362-380 CGCAGUUGGGCACUU 479UCAAAAGUGCCCAACUG 480 UUGATT CGTT 363 363-381 GCAGUUGGGCACUUU 481UUCAAAAGUGCCCAACU 482 UGAATT GCTT 364 364-382 CAGUUGGGCACUUUU 483CUUCAAAAGUGCCCAAC 484 GAAGTT UGTT 365 365-383 AGUUGGGCACUUUUG 485UCUUCAAAAGUGCCCAA 486 AAGATT CUTT 366 366-384 GUUGGGCACUUUUGA 487AUCUUCAAAAGUGCCCA 488 AGAUTT ACTT 367 367-385 UUGGGCACUUUUGAA 489GAUCUUCAAAAGUGCCC 490 GAUCTT AATT 368 368-386 UGGGCACUUUUGAAG 491UGAUCUUCAAAAGUGCC 492 AUCATT CATT 369 369-387 GGGCACUUUUGAAGA 493AUGAUCUUCAAAAGUG 494 UCAUTT CCCTT 377 377-395 UUGAAGAUCAUUUUC 495CUGAGAAAAUGAUCUU 496 UCAGTT CAATT 379 379-397 GAAGAUCAUUUUCUC 497GGCUGAGAAAAUGAUC 498 AGCCTT UUCTT 380 380-398 AAGAUCAUUUUCUCA 499AGGCUGAGAAAAUGAU 500 GCCUTT CUUTT 385 385-403 CAUUUUCUCAGCCUCC 501UCUGGAGGCUGAGAAA 502 AGATT AUGTT 394 394-412 AGCCUCCAGAGGAUG 503UGAACAUCCUCUGGAGG 504 UUCATT CUTT 396 396-414 CCUCCAGAGGAUGUUC 505AUUGAACAUCCUCUGGA 506 AAUTT GGTT 397 397-415 CUCCAGAGGAUGUUC 507UAUUGAACAUCCUCUGG 508 AAUATT AGTT 401 401-419 AGAGGAUGUUCAAUA 509CAGUUAUUGAACAUCCU 510 ACUGTT CUTT 403 403-421 AGGAUGUUCAAUAAC 511CACAGUUAUUGAACAUC 512 UGUGTT CUTT 407 407-425 UGUUCAAUAACUGUG 513ACCUCACAGUUAUUGAA 514 AGGUTT CATT 409 409-427 UUCAAUAACUGUGAG 515CCACCUCACAGUUAUUG 516 GUGGTT AATT 410 410-428 UCAAUAACUGUGAGG 517ACCACCUCACAGUUAUU 518 UGGUTT GATT 411 411-429 CAAUAACUGUGAGGU 519GACCACCUCACAGUUAU 520 GGUCTT UGTT 412 412-430 AAUAACUGUGAGGUG 521GGACCACCUCACAGUUA 522 GUCCTT UUTT 413 413-431 AUAACUGUGAGGUGG 523AGGACCACCUCACAGUU 524 UCCUTT AUTT 414 414-432 UAACUGUGAGGUGGU 525AAGGACCACCUCACAGU 526 CCUUTT UATT 416 416-434 ACUGUGAGGUGGUCC 527CCAAGGACCACCUCACA 528 UUGGTT GUTT 418 418-436 UGUGAGGUGGUCCUU 529UCCCAAGGACCACCUCA 530 GGGATT CATT 419 419-437 GUGAGGUGGUCCUUG 531UUCCCAAGGACCACCUC 532 GGAATT ACTT 425 425-443 UGGUCCUUGGGAAUU 533UCCAAAUUCCCAAGGAC 534 UGGATT CATT 431 431-449 UUGGGAAUUUGGAAA 535GUAAUUUCCAAAUUCCC 536 UUACTT AATT 432 432-450 UGGGAAUUUGGAAAU 537GGUAAUUUCCAAAUUCC 538 UACCTT CATT 433 433-451 GGGAAUUUGGAAAUU 539AGGUAAUUUCCAAAUU 540 ACCUTT CCCTT 434 434-452 GGAAUUUGGAAAUUA 541UAGGUAAUUUCCAAAU 542 CCUATT UCCTT 458 458-476 AGAGGAAUUAUGAUC 543GAAAGAUCAUAAUUCC 544 UUUCTT UCUTT 459 459-477 GAGGAAUUAUGAUCU 545GGAAAGAUCAUAAUUC 546 UUCCTT CUCTT 463 463-481 AAUUAUGAUCUUUCC 547AGAAGGAAAGAUCAUA 548 UUCUTT AUUTT 464 464-482 AUUAUGAUCUUUCCU 549AAGAAGGAAAGAUCAU 550 UCUUTT AAUTT 466 466-484 UAUGAUCUUUCCUUC 551UUAAGAAGGAAAGAUC 552 UUAATT AUATT 468 468-486 UGAUCUUUCCUUCUU 553CUUUAAGAAGGAAAGA 554 AAAGTT UCATT 471 471-489 UCUUUCCUUCUUAAA 555GGUCUUUAAGAAGGAA 556 GACCTT AGATT 476 476-494 CCUUCUUAAAGACCAU 557UGGAUGGUCUUUAAGA 558 CCATT AGGTT 477 477-495 CUUCUUAAAGACCAUC 559CUGGAUGGUCUUUAAG 560 CAGTT AAGTT 479 479-497 UCUUAAAGACCAUCCA 561UCCUGGAUGGUCUUUA 562 GGATT AGATT 481 481-499 UUAAAGACCAUCCAG 563CCUCCUGGAUGGUCUUU 564 GAGGTT AATT 482 482-500 UAAAGACCAUCCAGG 565ACCUCCUGGAUGGUCUU 566 AGGUTT UATT 492 492-510 CCAGGAGGUGGCUGG 567AUAACCAGCCACCUCCU 568 UUAUTT GGTT 493 493-511 CAGGAGGUGGCUGGU 569CAUAACCAGCCACCUCC 570 UAUGTT UGTT 494 494-512 AGGAGGUGGCUGGUU 571ACAUAACCAGCCACCUC 572 AUGUTT CUTT 495 495-513 GGAGGUGGCUGGUUA 573GACAUAACCAGCCACCU 574 UGUCTT CCTT 496 496-514 GAGGUGGCUGGUUAU 575GGACAUAACCAGCCACC 576 GUCCTT UCTT 497 497-515 AGGUGGCUGGUUAUG 577AGGACAUAACCAGCCAC 578 UCCUTT CUTT 499 499-517 GUGGCUGGUUAUGUC 579UGAGGACAUAACCAGCC 580 CUCATT ACTT 520 520-538 GCCCUCAACACAGUGG 581GCUCCACUGUGUUGAGG 582 AGCTT GCTT 542 542-560 UUCCUUUGGAAAACC 583UGCAGGUUUUCCAAAG 584 UGCATT GAATT 543 543-561 UCCUUUGGAAAACCU 585CUGCAGGUUUUCCAAAG 586 GCAGTT GATT 550 550-568 GAAAACCUGCAGAUC 587UGAUGAUCUGCAGGUU 588 AUCATT UUCTT 551 551-569 AAAACCUGCAGAUCA 589CUGAUGAUCUGCAGGU 590 UCAGTT UUUTT 553 553-571 AACCUGCAGAUCAUCA 591CUCUGAUGAUCUGCAGG 592 GAGTT UUTT 556 556-574 CUGCAGAUCAUCAGA 593UUCCUCUGAUGAUCUGC 594 GGAATT AGTT 586 586-604 GAAAAUUCCUAUGCC 595CUAAGGCAUAGGAAUU 596 UUAGTT UUCTT 587 587-605 AAAAUUCCUAUGCCU 597GCUAAGGCAUAGGAAU 598 UAGCTT UUUTT 589 589-607 AAUUCCUAUGCCUUA 599CUGCUAAGGCAUAGGA 600 GCAGTT AUUTT 592 592-610 UCCUAUGCCUUAGCAG 601AGACUGCUAAGGCAUA 602 UCUTT GGATT 593 593-611 CCUAUGCCUUAGCAGU 603AAGACUGCUAAGGCAU 604 CUUTT AGGTT 594 594-612 CUAUGCCUUAGCAGUC 605UAAGACUGCUAAGGCA 606 UUATT UAGTT 596 596-614 AUGCCUUAGCAGUCU 607GAUAAGACUGCUAAGG 608 UAUCTT CAUTT 597 597-615 UGCCUUAGCAGUCUU 609AGAUAAGACUGCUAAG 610 AUCUTT GCATT 598 598-616 GCCUUAGCAGUCUUA 611UAGAUAAGACUGCUAA 612 UCUATT GGCTT 599 599-617 CCUUAGCAGUCUUAUC 613UUAGAUAAGACUGCUA 614 UAATT AGGTT 600 600-618 CUUAGCAGUCUUAUC 615GUUAGAUAAGACUGCU 616 UAACTT AAGTT 601 601-619 UUAGCAGUCUUAUCU 617AGUUAGAUAAGACUGC 618 AACUTT UAATT 602 602-620 UAGCAGUCUUAUCUA 619UAGUUAGAUAAGACUG 620 ACUATT CUATT 603 603-621 AGCAGUCUUAUCUAA 621AUAGUUAGAUAAGACU 622 CUAUTT GCUTT 604 604-622 GCAGUCUUAUCUAAC 623CAUAGUUAGAUAAGAC 624 UAUGTT UGCTT 605 605-623 CAGUCUUAUCUAACU 625UCAUAGUUAGAUAAGA 626 AUGATT CUGTT 608 608-626 UCUUAUCUAACUAUG 627GCAUCAUAGUUAGAUA 628 AUGCTT AGATT 609 609-627 CUUAUCUAACUAUGA 629UGCAUCAUAGUUAGAU 630 UGCATT AAGTT 610 610-628 UUAUCUAACUAUGAU 631UUGCAUCAUAGUUAGA 632 GCAATT UAATT 611 611-629 UAUCUAACUAUGAUG 633UUUGCAUCAUAGUUAG 634 CAAATT AUATT 612 612-630 AUCUAACUAUGAUGC 635AUUUGCAUCAUAGUUA 636 AAAUTT GAUTT 613 613-631 UCUAACUAUGAUGCA 637UAUUUGCAUCAUAGUU 638 AAUATT AGATT 614 614-632 CUAACUAUGAUGCAA 639UUAUUUGCAUCAUAGU 640 AUAATT UAGTT 616 616-634 AACUAUGAUGCAAAU 641UUUUAUUUGCAUCAUA 642 AAAATT GUUTT 622 622-640 GAUGCAAAUAAAACC 643GUCCGGUUUUAUUUGC 644 GGACTT AUCTT 623 623-641 AUGCAAAUAAAACCG 645AGUCCGGUUUUAUUUG 646 GACUTT CAUTT 624 624-642 UGCAAAUAAAACCGG 647CAGUCCGGUUUUAUUU 648 ACUGTT GCATT 626 626-644 CAAAUAAAACCGGAC 649UUCAGUCCGGUUUUAU 650 UGAATT UUGTT 627 627-645 AAAUAAAACCGGACU 651CUUCAGUCCGGUUUUAU 652 GAAGTT UUTT 628 628-646 AAUAAAACCGGACUG 653CCUUCAGUCCGGUUUUA 654 AAGGTT UUTT 630 630-648 UAAAACCGGACUGAA 655CUCCUUCAGUCCGGUUU 656 GGAGTT UATT 631 631-649 AAAACCGGACUGAAG 657GCUCCUUCAGUCCGGUU 658 GAGCTT UUTT 632 632-650 AAACCGGACUGAAGG 659AGCUCCUUCAGUCCGGU 660 AGCUTT UUTT 633 633-651 AACCGGACUGAAGGA 661CAGCUCCUUCAGUCCGG 662 GCUGTT UUTT 644 644-662 AGGAGCUGCCCAUGA 663UUUCUCAUGGGCAGCUC 664 GAAATT CUTT 665 665-683 UACAGGAAAUCCUGC 665CCAUGCAGGAUUUCCUG 666 AUGGTT UATT 668 668-686 AGGAAAUCCUGCAUG 667GCGCCAUGCAGGAUUUC 668 GCGCTT CUTT 669 669-687 GGAAAUCCUGCAUGG 669GGCGCCAUGCAGGAUUU 670 CGCCTT CCTT 670 670-688 GAAAUCCUGCAUGGC 671CGGCGCCAUGCAGGAUU 672 GCCGTT UCTT 671 671-689 AAAUCCUGCAUGGCGC 673ACGGCGCCAUGCAGGAU 674 CGUTT UUTT 672 672-690 AAUCCUGCAUGGCGCC 675CACGGCGCCAUGCAGGA 676 GUGTT UUTT 674 674-692 UCCUGCAUGGCGCCGU 677CGCACGGCGCCAUGCAG 678 GCGTT GATT 676 676-694 CUGCAUGGCGCCGUGC 679ACCGCACGGCGCCAUGC 680 GGUTT AGTT 677 677-695 UGCAUGGCGCCGUGCG 681AACCGCACGGCGCCAUG 682 GUUTT CATT 678 678-696 GCAUGGCGCCGUGCGG 683GAACCGCACGGCGCCAU 684 UUCTT GCTT 680 680-698 AUGGCGCCGUGCGGU 685CUGAACCGCACGGCGCC 686 UCAGTT AUTT 681 681-699 UGGCGCCGUGCGGUUC 687GCUGAACCGCACGGCGC 688 AGCTT CATT 682 682-700 GGCGCCGUGCGGUUCA 689UGCUGAACCGCACGGCG 690 GCATT CCTT 683 683-701 GCGCCGUGCGGUUCAG 691UUGCUGAACCGCACGGC 692 CAATT GCTT 684 684-702 CGCCGUGCGGUUCAGC 693GUUGCUGAACCGCACGG 694 AACTT CGTT 685 685-703 GCCGUGCGGUUCAGCA 695UGUUGCUGAACCGCACG 696 ACATT GCTT 686 686-704 CCGUGCGGUUCAGCAA 697UUGUUGCUGAACCGCAC 698 CAATT GGTT 688 688-706 GUGCGGUUCAGCAAC 699GGUUGUUGCUGAACCGC 700 AACCTT ACTT 690 690-708 GCGGUUCAGCAACAAC 701AGGGUUGUUGCUGAAC 702 CCUTT CGCTT 692 692-710 GGUUCAGCAACAACCC 703GCAGGGUUGUUGCUGA 704 UGCTT ACCTT 698 698-716 GCAACAACCCUGCCCU 705CACAGGGCAGGGUUGU 706 GUGTT UGCTT 700 700-718 AACAACCCUGCCCUGU 707UGCACAGGGCAGGGUU 708 GCATT GUUTT 719 719-737 ACGUGGAGAGCAUCC 709CACUGGAUGCUCUCCAC 710 AGUGTT GUTT 720 720-738 CGUGGAGAGCAUCCA 711CCACUGGAUGCUCUCCA 712 GUGGTT CGTT 721 721-739 GUGGAGAGCAUCCAG 713GCCACUGGAUGCUCUCC 714 UGGCTT ACTT 724 724-742 GAGAGCAUCCAGUGG 715CCCGCCACUGGAUGCUC 716 CGGGTT UCTT 725 725-743 AGAGCAUCCAGUGGC 717UCCCGCCACUGGAUGCU 718 GGGATT CUTT 726 726-744 GAGCAUCCAGUGGCG 719GUCCCGCCACUGGAUGC 720 GGACTT UCTT 733 733-751 CAGUGGCGGGACAUA 721UGACUAUGUCCCGCCAC 722 GUCATT UGTT 734 734-752 AGUGGCGGGACAUAG 723CUGACUAUGUCCCGCCA 724 UCAGTT CUTT 736 736-754 UGGCGGGACAUAGUC 725UGCUGACUAUGUCCCGC 726 AGCATT CATT 737 737-755 GGCGGGACAUAGUCA 727CUGCUGACUAUGUCCCG 728 GCAGTT CCTT 763 763-781 CUCAGCAACAUGUCGA 729CCAUCGACAUGUUGCUG 730 UGGTT AGTT 765 765-783 CAGCAACAUGUCGAU 731GUCCAUCGACAUGUUGC 732 GGACTT UGTT 766 766-784 AGCAACAUGUCGAUG 733AGUCCAUCGACAUGUUG 734 GACUTT CUTT 767 767-785 GCAACAUGUCGAUGG 735AAGUCCAUCGACAUGUU 736 ACUUTT GCTT 769 769-787 AACAUGUCGAUGGAC 737GGAAGUCCAUCGACAUG 738 UUCCTT UUTT 770 770-788 ACAUGUCGAUGGACU 739UGGAAGUCCAUCGACAU 740 UCCATT GUTT 771 771-789 CAUGUCGAUGGACUU 741CUGGAAGUCCAUCGACA 742 CCAGTT UGTT 772 772-790 AUGUCGAUGGACUUC 743UCUGGAAGUCCAUCGAC 744 CAGATT AUTT 775 775-793 UCGAUGGACUUCCAG 745GGUUCUGGAAGUCCAUC 746 AACCTT GATT 789 789-807 GAACCACCUGGGCAGC 747GCAGCUGCCCAGGUGGU 748 UGCTT UCTT 798 798-816 GGGCAGCUGCCAAAA 749ACACUUUUGGCAGCUGC 750 GUGUTT CCTT 800 800-818 GCAGCUGCCAAAAGU 751UCACACUUUUGGCAGCU 752 GUGATT GCTT 805 805-823 UGCCAAAAGUGUGAU 753UUGGAUCACACUUUUG 754 CCAATT GCATT 806 806-824 GCCAAAAGUGUGAUC 755CUUGGAUCACACUUUUG 756 CAAGTT GCTT 807 807-825 CCAAAAGUGUGAUCC 757GCUUGGAUCACACUUUU 758 AAGCTT GGTT 810 810-828 AAAGUGUGAUCCAAG 759ACAGCUUGGAUCACACU 760 CUGUTT UUTT 814 814-832 UGUGAUCCAAGCUGU 761UGGGACAGCUUGGAUC 762 CCCATT ACATT 815 815-833 GUGAUCCAAGCUGUCC 763UUGGGACAGCUUGGAU 764 CAATT CACTT 817 817-835 GAUCCAAGCUGUCCCA 765CAUUGGGACAGCUUGG 766 AUGTT AUCTT 818 818-836 AUCCAAGCUGUCCCAA 767CCAUUGGGACAGCUUGG 768 UGGTT AUTT 819 819-837 UCCAAGCUGUCCCAAU 769CCCAUUGGGACAGCUUG 770 GGGTT GATT 820 820-838 CCAAGCUGUCCCAAUG 771UCCCAUUGGGACAGCUU 772 GGATT GGTT 821 821-839 CAAGCUGUCCCAAUGG 773CUCCCAUUGGGACAGCU 774 GAGTT UGTT 823 823-841 AGCUGUCCCAAUGGG 775AGCUCCCAUUGGGACAG 776 AGCUTT CUTT 826 826-844 UGUCCCAAUGGGAGC 777AGCAGCUCCCAUUGGGA 778 UGCUTT CATT 847 847-865 GGUGCAGGAGAGGAG 779AGUUCUCCUCUCCUGCA 780 AACUTT CCTT 871 871-889 AAACUGACCAAAAUC 781AGAUGAUUUUGGUCAG 782 AUCUTT UUUTT 872 872-890 AACUGACCAAAAUCA 783CAGAUGAUUUUGGUCA 784 UCUGTT GUUTT 873 873-891 ACUGACCAAAAUCAUC 785ACAGAUGAUUUUGGUC 786 UGUTT AGUTT 877 877-895 ACCAAAAUCAUCUGU 787GGGCACAGAUGAUUUU 788 GCCCTT GGUTT 878 878-896 CCAAAAUCAUCUGUGC 789UGGGCACAGAUGAUUU 790 CCATT UGGTT 881 881-899 AAAUCAUCUGUGCCCA 791UGCUGGGCACAGAUGA 792 GCATT UUUTT 890 890-908 GUGCCCAGCAGUGCUC 793CCGGAGCACUGCUGGGC 794 CGGTT ACTT 892 892-910 GCCCAGCAGUGCUCCG 795GCCCGGAGCACUGCUGG 796 GGCTT GCTT 929 929-947 CCAGUGACUGCUGCCA 797UUGUGGCAGCAGUCACU 798 CAATT GGTT 930 930-948 CAGUGACUGCUGCCAC 799GUUGUGGCAGCAGUCAC 800 AACTT UGTT 979 979-997 GAGAGCGACUGCCUG 801AGACCAGGCAGUCGCUC 802 GUCUTT UCTT 980 980-998 AGAGCGACUGCCUGG 803CAGACCAGGCAGUCGCU 804 UCUGTT CUTT 981 981-999 GAGCGACUGCCUGGUC 805GCAGACCAGGCAGUCGC 806 UGCTT UCTT 982  982-1000 AGCGACUGCCUGGUCU 807GGCAGACCAGGCAGUCG 808 GCCTT CUTT 983  983-1001 GCGACUGCCUGGUCUG 809CGGCAGACCAGGCAGUC 810 CCGTT GCTT 984  984-1002 CGACUGCCUGGUCUGC 811GCGGCAGACCAGGCAGU 812 CGCTT CGTT 989  989-1007 GCCUGGUCUGCCGCAA 813AAUUUGCGGCAGACCAG 814 AUUTT GCTT 990  990-1008 CCUGGUCUGCCGCAAA 815GAAUUUGCGGCAGACCA 816 UUCTT GGTT 991  991-1009 CUGGUCUGCCGCAAAU 817GGAAUUUGCGGCAGACC 818 UCCTT AGTT 992  992-1010 UGGUCUGCCGCAAAU 819CGGAAUUUGCGGCAGAC 820 UCCGTT CATT 994  994-1012 GUCUGCCGCAAAUUCC 821CUCGGAAUUUGCGGCAG 822 GAGTT ACTT 995  995-1013 UCUGCCGCAAAUUCCG 823UCUCGGAAUUUGCGGCA 824 AGATT GATT 996  996-1014 CUGCCGCAAAUUCCGA 825GUCUCGGAAUUUGCGGC 826 GACTT AGTT 997  997-1015 UGCCGCAAAUUCCGAG 827CGUCUCGGAAUUUGCGG 828 ACGTT CATT 999  999-1017 CCGCAAAUUCCGAGAC 829UUCGUCUCGGAAUUUGC 830 GAATT GGTT 1004 1004-1022 AAUUCCGAGACGAAG 831GUGGCUUCGUCUCGGAA 832 CCACTT UUTT 1005 1005-1023 AUUCCGAGACGAAGCC 833CGUGGCUUCGUCUCGGA 834 ACGTT AUTT 1006 1006-1024 UUCCGAGACGAAGCCA 835ACGUGGCUUCGUCUCGG 836 CGUTT AATT 1007 1007-1025 UCCGAGACGAAGCCAC 837CACGUGGCUUCGUCUCG 838 GUGTT GATT 1008 1008-1026 CCGAGACGAAGCCACG 839GCACGUGGCUUCGUCUC 840 UGCTT GGTT 1010 1010-1028 GAGACGAAGCCACGU 841UUGCACGUGGCUUCGUC 842 GCAATT UCTT 1013 1013-1031 ACGAAGCCACGUGCAA 843UCCUUGCACGUGGCUUC 844 GGATT GUTT 1014 1014-1032 CGAAGCCACGUGCAAG 845GUCCUUGCACGUGGCUU 846 GACTT CGTT 1015 1015-1033 GAAGCCACGUGCAAG 847UGUCCUUGCACGUGGCU 848 GACATT UCTT 1016 1016-1034 AAGCCACGUGCAAGG 849GUGUCCUUGCACGUGGC 850 ACACTT UUTT 1040 1040-1058 CCCCACUCAUGCUCUA 851UUGUAGAGCAUGAGUG 852 CAATT GGGTT 1042 1042-1060 CCACUCAUGCUCUACA 853GGUUGUAGAGCAUGAG 854 ACCTT UGGTT 1044 1044-1062 ACUCAUGCUCUACAAC 855GGGGUUGUAGAGCAUG 856 CCCTT AGUTT 1047 1047-1065 CAUGCUCUACAACCCC 857GGUGGGGUUGUAGAGC 858 ACCTT AUGTT 1071 1071-1089 CCAGAUGGAUGUGAA 859GGGGUUCACAUCCAUCU 860 CCCCTT GGTT 1073 1073-1091 AGAUGGAUGUGAACC 861UCGGGGUUCACAUCCAU 862 CCGATT CUTT 1074 1074-1092 GAUGGAUGUGAACCC 863CUCGGGGUUCACAUCCA 864 CGAGTT UCTT 1075 1075-1093 AUGGAUGUGAACCCC 865CCUCGGGGUUCACAUCC 866 GAGGTT AUTT 1077 1077-1095 GGAUGUGAACCCCGA 867GCCCUCGGGGUUCACAU 868 GGGCTT CCTT 1078 1078-1096 GAUGUGAACCCCGAG 869UGCCCUCGGGGUUCACA 870 GGCATT UCTT 1080 1080-1098 UGUGAACCCCGAGGGC 871UUUGCCCUCGGGGUUCA 872 AAATT CATT 1084 1084-1102 AACCCCGAGGGCAAAU 873UGUAUUUGCCCUCGGGG 874 ACATT UUTT 1085 1085-1103 ACCCCGAGGGCAAAUA 875CUGUAUUUGCCCUCGGG 876 CAGTT GUTT 1087 1087-1105 CCCGAGGGCAAAUACA 877AGCUGUAUUUGCCCUCG 878 GCUTT GGTT 1088 1088-1106 CCGAGGGCAAAUACA 879AAGCUGUAUUUGCCCUC 880 GCUUTT GGTT 1089 1089-1107 CGAGGGCAAAUACAG 881AAAGCUGUAUUUGCCCU 882 CUUUTT CGTT 1096 1096-1114 AAAUACAGCUUUGGU 883UGGCACCAAAGCUGUAU 884 GCCATT UUTT 1097 1097-1115 AAUACAGCUUUGGUG 885GUGGCACCAAAGCUGUA 886 CCACTT UUTT 1098 1098-1116 AUACAGCUUUGGUGC 887GGUGGCACCAAAGCUGU 888 CACCTT AUTT 1104 1104-1122 CUUUGGUGCCACCUGC 889CACGCAGGUGGCACCAA 890 GUGTT AGTT 1106 1106-1124 UUGGUGCCACCUGCGU 891UUCACGCAGGUGGCACC 892 GAATT AATT 1112 1112-1130 CCACCUGCGUGAAGAA 893CACUUCUUCACGCAGGU 894 GUGTT GGTT 1116 1116-1134 CUGCGUGAAGAAGUG 895GGGACACUUCUUCACGC 896 UCCCTT AGTT 1117 1117-1135 UGCGUGAAGAAGUGU 897GGGGACACUUCUUCACG 898 CCCCTT CATT 1118 1118-1136 GCGUGAAGAAGUGUC 899CGGGGACACUUCUUCAC 900 CCCGTT GCTT 1119 1119-1137 CGUGAAGAAGUGUCC 901ACGGGGACACUUCUUCA 902 CCGUTT CGTT 1120 1120-1138 GUGAAGAAGUGUCCC 903UACGGGGACACUUCUUC 904 CGUATT ACTT 1121 1121-1139 UGAAGAAGUGUCCCC 905UUACGGGGACACUUCUU 906 GUAATT CATT 1122 1122-1140 GAAGAAGUGUCCCCG 907AUUACGGGGACACUUCU 908 UAAUTT UCTT 1123 1123-1141 AAGAAGUGUCCCCGU 909AAUUACGGGGACACUUC 910 AAUUTT UUTT 1124 1124-1142 AGAAGUGUCCCCGUA 911UAAUUACGGGGACACU 912 AUUATT UCUTT 1125 1125-1143 GAAGUGUCCCCGUAA 913AUAAUUACGGGGACAC 914 UUAUTT UUCTT 1126 1126-1144 AAGUGUCCCCGUAAU 915CAUAAUUACGGGGACAC 916 UAUGTT UUTT 1127 1127-1145 AGUGUCCCCGUAAUU 917ACAUAAUUACGGGGAC 918 AUGUTT ACUTT 1128 1128-1146 GUGUCCCCGUAAUUA 919CACAUAAUUACGGGGAC 920 UGUGTT ACTT 1129 1129-1147 UGUCCCCGUAAUUAU 921CCACAUAAUUACGGGGA 922 GUGGTT CATT 1130 1130-1148 GUCCCCGUAAUUAUG 923ACCACAUAAUUACGGGG 924 UGGUTT ACTT 1132 1132-1150 CCCCGUAAUUAUGUG 925UCACCACAUAAUUACGG 926 GUGATT GGTT 1134 1134-1152 CCGUAAUUAUGUGGU 927UGUCACCACAUAAUUAC 928 GACATT GGTT 1136 1136-1154 GUAAUUAUGUGGUGA 929UCUGUCACCACAUAAUU 930 CAGATT ACTT 1137 1137-1155 UAAUUAUGUGGUGAC 931AUCUGUCACCACAUAAU 932 AGAUTT UATT 1138 1138-1156 AAUUAUGUGGUGACA 933GAUCUGUCACCACAUAA 934 GAUCTT UUTT 1139 1139-1157 AUUAUGUGGUGACAG 935UGAUCUGUCACCACAUA 936 AUCATT AUTT 1140 1140-1158 UUAUGUGGUGACAGA 937GUGAUCUGUCACCACAU 938 UCACTT AATT 1142 1142-1160 AUGUGGUGACAGAUC 939CCGUGAUCUGUCACCAC 940 ACGGTT AUTT 1145 1145-1163 UGGUGACAGAUCACG 941GAGCCGUGAUCUGUCAC 942 GCUCTT CATT 1147 1147-1165 GUGACAGAUCACGGC 943ACGAGCCGUGAUCUGUC 944 UCGUTT ACTT 1148 1148-1166 UGACAGAUCACGGCUC 945CACGAGCCGUGAUCUGU 946 GUGTT CATT 1149 1149-1167 GACAGAUCACGGCUCG 947GCACGAGCCGUGAUCUG 948 UGCTT UCTT 1150 1150-1168 ACAGAUCACGGCUCGU 949CGCACGAGCCGUGAUCU 950 GCGTT GUTT 1151 1151-1169 CAGAUCACGGCUCGUG 951ACGCACGAGCCGUGAUC 952 CGUTT UGTT 1152 1152-1170 AGAUCACGGCUCGUGC 953GACGCACGAGCCGUGAU 954 GUCTT CUTT 1153 1153-1171 GAUCACGGCUCGUGCG 955GGACGCACGAGCCGUGA 956 UCCTT UCTT 1154 1154-1172 AUCACGGCUCGUGCGU 957CGGACGCACGAGCCGUG 958 CCGTT AUTT 1155 1155-1173 UCACGGCUCGUGCGUC 959UCGGACGCACGAGCCGU 960 CGATT GATT 1156 1156-1174 CACGGCUCGUGCGUCC 961CUCGGACGCACGAGCCG 962 GAGTT UGTT 1157 1157-1175 ACGGCUCGUGCGUCCG 963GCUCGGACGCACGAGCC 964 AGCTT GUTT 1160 1160-1178 GCUCGUGCGUCCGAGC 965CAGGCUCGGACGCACGA 966 CUGTT GCTT 1200 1200-1218 GGAGGAAGACGGCGU 967GCGGACGCCGUCUUCCU 968 CCGCTT CCTT 1201 1201-1219 GAGGAAGACGGCGUC 969UGCGGACGCCGUCUUCC 970 CGCATT UCTT 1203 1203-1221 GGAAGACGGCGUCCGC 971CUUGCGGACGCCGUCUU 972 AAGTT CCTT 1204 1204-1222 GAAGACGGCGUCCGCA 973ACUUGCGGACGCCGUCU 974 AGUTT UCTT 1205 1205-1223 AAGACGGCGUCCGCAA 975CACUUGCGGACGCCGUC 976 GUGTT UUTT 1207 1207-1225 GACGGCGUCCGCAAGU 977UACACUUGCGGACGCCG 978 GUATT UCTT 1208 1208-1226 ACGGCGUCCGCAAGUG 979UUACACUUGCGGACGCC 980 UAATT GUTT 1211 1211-1229 GCGUCCGCAAGUGUA 981UUCUUACACUUGCGGAC 982 AGAATT GCTT 1212 1212-1230 CGUCCGCAAGUGUAA 983CUUCUUACACUUGCGGA 984 GAAGTT CGTT 1213 1213-1231 GUCCGCAAGUGUAAG 985ACUUCUUACACUUGCGG 986 AAGUTT ACTT 1214 1214-1232 UCCGCAAGUGUAAGA 987CACUUCUUACACUUGCG 988 AGUGTT GATT 1215 1215-1233 CCGCAAGUGUAAGAA 989GCACUUCUUACACUUGC 990 GUGCTT GGTT 1216 1216-1234 CGCAAGUGUAAGAAG 991CGCACUUCUUACACUUG 992 UGCGTT CGTT 1217 1217-1235 GCAAGUGUAAGAAGU 993UCGCACUUCUUACACUU 994 GCGATT GCTT 1219 1219-1237 AAGUGUAAGAAGUGC 995CUUCGCACUUCUUACAC 996 GAAGTT UUTT 1220 1220-1238 AGUGUAAGAAGUGCG 997CCUUCGCACUUCUUACA 998 AAGGTT CUTT 1221 1221-1239 GUGUAAGAAGUGCGA 999CCCUUCGCACUUCUUAC 1000 AGGGTT ACTT 1222 1222-1240 UGUAAGAAGUGCGAA 1001GCCCUUCGCACUUCUUA 1002 GGGCTT CATT 1223 1223-1241 GUAAGAAGUGCGAAG 1003GGCCCUUCGCACUUCUU 1004 GGCCTT ACTT 1224 1224-1242 UAAGAAGUGCGAAGG 1005AGGCCCUUCGCACUUCU 1006 GCCUTT UATT 1225 1225-1243 AAGAAGUGCGAAGGG 1007AAGGCCCUUCGCACUUC 1008 CCUUTT UUTT 1226 1226-1244 AGAAGUGCGAAGGGC 1009CAAGGCCCUUCGCACUU 1010 CUUGTT CUTT 1229 1229-1247 AGUGCGAAGGGCCUU 1011CGGCAAGGCCCUUCGCA 1012 GCCGTT CUTT 1230 1230-1248 GUGCGAAGGGCCUUG 1013GCGGCAAGGCCCUUCGC 1014 CCGCTT ACTT 1231 1231-1249 UGCGAAGGGCCUUGCC 1015UGCGGCAAGGCCCUUCG 1016 GCATT CATT 1232 1232-1250 GCGAAGGGCCUUGCCG 1017UUGCGGCAAGGCCCUUC 1018 CAATT GCTT 1233 1233-1251 CGAAGGGCCUUGCCGC 1019UUUGCGGCAAGGCCCUU 1020 AAATT CGTT 1235 1235-1253 AAGGGCCUUGCCGCAA 1021ACUUUGCGGCAAGGCCC 1022 AGUTT UUTT 1236 1236-1254 AGGGCCUUGCCGCAAA 1023CACUUUGCGGCAAGGCC 1024 GUGTT CUTT 1237 1237-1255 GGGCCUUGCCGCAAAG 1025ACACUUUGCGGCAAGGC 1026 UGUTT CCTT 1238 1238-1256 GGCCUUGCCGCAAAGU 1027CACACUUUGCGGCAAGG 1028 GUGTT CCTT 1239 1239-1257 GCCUUGCCGCAAAGUG 1029ACACACUUUGCGGCAAG 1030 UGUTT GCTT 1241 1241-1259 CUUGCCGCAAAGUGU 1031UUACACACUUUGCGGCA 1032 GUAATT AGTT 1261 1261-1279 GGAAUAGGUAUUGGU 1033AUUCACCAAUACCUAUU 1034 GAAUTT CCTT 1262 1262-1280 GAAUAGGUAUUGGUG 1035AAUUCACCAAUACCUAU 1036 AAUUTT UCTT 1263 1263-1281 AAUAGGUAUUGGUGA 1037AAAUUCACCAAUACCUA 1038 AUUUTT UUTT 1264 1264-1282 AUAGGUAUUGGUGAA 1039UAAAUUCACCAAUACCU 1040 UUUATT AUTT 1266 1266-1284 AGGUAUUGGUGAAUU 1041UUUAAAUUCACCAAUAC 1042 UAAATT CUTT 1267 1267-1285 GGUAUUGGUGAAUUU 1043CUUUAAAUUCACCAAUA 1044 AAAGTT CCTT 1289 1289-1307 CACUCUCCAUAAAUGC 1045GUAGCAUUUAUGGAGA 1046 UACTT GUGTT 1313 1313-1331 UUAAACACUUCAAAA 1047CAGUUUUUGAAGUGUU 1048 ACUGTT UAATT 1320 1320-1338 CUUCAAAAACUGCACC 1049GGAGGUGCAGUUUUUG 1050 UCCTT AAGTT 1321 1321-1339 UUCAAAAACUGCACCU 1051UGGAGGUGCAGUUUUU 1052 CCATT GAATT 1322 1322-1340 UCAAAAACUGCACCUC 1053AUGGAGGUGCAGUUUU 1054 CAUTT UGATT 1323 1323-1341 CAAAAACUGCACCUCC 1055GAUGGAGGUGCAGUUU 1056 AUCTT UUGTT 1324 1324-1342 AAAAACUGCACCUCCA 1057UGAUGGAGGUGCAGUU 1058 UCATT UUUTT 1328 1328-1346 ACUGCACCUCCAUCAG 1059CCACUGAUGGAGGUGCA 1060 UGGTT GUTT 1332 1332-1350 CACCUCCAUCAGUGGC 1061AUCGCCACUGAUGGAGG 1062 GAUTT UGTT 1333 1333-1351 ACCUCCAUCAGUGGCG 1063GAUCGCCACUGAUGGAG 1064 AUCTT GUTT 1335 1335-1353 CUCCAUCAGUGGCGAU 1065GAGAUCGCCACUGAUGG 1066 CUCTT AGTT 1338 1338-1356 CAUCAGUGGCGAUCUC 1067GUGGAGAUCGCCACUGA 1068 CACTT UGTT 1344 1344-1362 UGGCGAUCUCCACAUC 1069CAGGAUGUGGAGAUCG 1070 CUGTT CCATT 1345 1345-1363 GGCGAUCUCCACAUCC 1071GCAGGAUGUGGAGAUC 1072 UGCTT GCCTT 1346 1346-1364 GCGAUCUCCACAUCCU 1073GGCAGGAUGUGGAGAU 1074 GCCTT CGCTT 1347 1347-1365 CGAUCUCCACAUCCUG 1075CGGCAGGAUGUGGAGA 1076 CCGTT UCGTT 1348 1348-1366 GAUCUCCACAUCCUGC 1077CCGGCAGGAUGUGGAG 1078 CGGTT AUCTT 1353 1353-1371 CCACAUCCUGCCGGUG 1079UGCCACCGGCAGGAUGU 1080 GCATT GGTT 1354 1354-1372 CACAUCCUGCCGGUGG 1081AUGCCACCGGCAGGAUG 1082 CAUTT UGTT 1355 1355-1373 ACAUCCUGCCGGUGGC 1083AAUGCCACCGGCAGGAU 1084 AUUTT GUTT 1357 1357-1375 AUCCUGCCGGUGGCAU 1085UAAAUGCCACCGGCAGG 1086 UUATT AUTT 1360 1360-1378 CUGCCGGUGGCAUUU 1087CCCUAAAUGCCACCGGC 1088 AGGGTT AGTT 1361 1361-1379 UGCCGGUGGCAUUUA 1089CCCCUAAAUGCCACCGG 1090 GGGGTT CATT 1362 1362-1380 GCCGGUGGCAUUUAG 1091ACCCCUAAAUGCCACCG 1092 GGGUTT GCTT 1363 1363-1381 CCGGUGGCAUUUAGG 1093CACCCCUAAAUGCCACC 1094 GGUGTT GGTT 1366 1366-1384 GUGGCAUUUAGGGGU 1095AGUCACCCCUAAAUGCC 1096 GACUTT ACTT 1369 1369-1387 GCAUUUAGGGGUGAC 1097AGGAGUCACCCCUAAAU 1098 UCCUTT GCTT 1370 1370-1388 CAUUUAGGGGUGACU 1099AAGGAGUCACCCCUAAA 1100 CCUUTT UGTT 1371 1371-1389 AUUUAGGGGUGACUC 1101GAAGGAGUCACCCCUAA 1102 CUUCTT AUTT 1372 1372-1390 UUUAGGGGUGACUCC 1103UGAAGGAGUCACCCCUA 1104 UUCATT AATT 1373 1373-1391 UUAGGGGUGACUCCU 1105GUGAAGGAGUCACCCCU 1106 UCACTT AATT 1374 1374-1392 UAGGGGUGACUCCUU 1107UGUGAAGGAGUCACCCC 1108 CACATT UATT 1404 1404-1422 UCUGGAUCCACAGGA 1109CAGUUCCUGUGGAUCCA 1110 ACUGTT GATT 1408 1408-1426 GAUCCACAGGAACUG 1111UAUCCAGUUCCUGUGGA 1112 GAUATT UCTT 1409 1409-1427 AUCCACAGGAACUGG 1113AUAUCCAGUUCCUGUGG 1114 AUAUTT AUTT 1411 1411-1429 CCACAGGAACUGGAU 1115GAAUAUCCAGUUCCUGU 1116 AUUCTT GGTT 1412 1412-1430 CACAGGAACUGGAUA 1117AGAAUAUCCAGUUCCUG 1118 UUCUTT UGTT 1419 1419-1437 ACUGGAUAUUCUGAA 1119GGUUUUCAGAAUAUCC 1120 AACCTT AGUTT 1426 1426-1444 AUUCUGAAAACCGUA 1121CCUUUACGGUUUUCAGA 1122 AAGGTT AUTT 1427 1427-1445 UUCUGAAAACCGUAA 1123UCCUUUACGGUUUUCAG 1124 AGGATT AATT 1430 1430-1448 UGAAAACCGUAAAGG 1125AUUUCCUUUACGGUUU 1126 AAAUTT UCATT 1431 1431-1449 GAAAACCGUAAAGGA 1127GAUUUCCUUUACGGUU 1128 AAUCTT UUCTT

TABLE 3 EGFR siRNA Sequences with Chemical Modifications Sequence SEQSEQ position in sense strand sequence (5′- ID antisense strand sequenceID hs Id # NM_005228.3 3′) NO: (5′-3′) NO: 68 68-86cgGfcCfgGfaGfuCfcCfgAf 1129 UfAfgCfuCfgGfgAfcUfcCfgG 1130 gCfuAfdTsdTfcCfgdTsdT 71 71-89 ccGfgAfgUfcCfcGfaGfcUf 1131 GfGfcUfaGfcUfcGfgGfaCfuC1132 aGfcCfdTsdT fcGfgdTsdT 72 72-90 cgGfaGfuCfcCfgAfgCfuAf 1133GfGfgCfuAfgCfuCfgGfgAfcU 1134 gCfcCfdTsdT fcCfgdTsdT 73 73-91ggAfgUfcCfcGfaGfcUfaGf 1135 GfGfgGfcUfaGfcUfcGfgGfaC 1136 cCfcCfdTsdTfuCfcdTsdT 74 74-92 gaGfuCfcCfgAfgCfuAfgCf 1137 CfGfgGfgCfuAfgCfuCfgGfgA1138 cCfcGfdTsdT fcUfcdTsdT 75 75-93 agUfcCfcGfaGfcUfaGfcCf 1139CfCfgGfgGfcUfaGfcUfcGfgG 1140 cCfgGfdTsdT faCfudTsdT 76 76-94guCfcCfgAfgCfuAfgCfcCf 1141 GfCfcGfgGfgCfuAfgCfuCfgG 1142 cGfgCfdTsdTfgAfcdTsdT 78 78-96 ccCfgAfgCfuAfgCfcCfcGf 1143 CfCfgCfcGfgGfgCfuAfgCfuC1144 gCfgGfdTsdT fgGfgdTsdT 114 114-132 ggAfcGfaCfaGfgCfcAfcCf 1145AfCfgAfgGfuGfgCfcUfgUfcG 1146 uCfgUfdTsdT fuCfcdTsdT 115 115-133gaCfgAfcAfgGfcCfaCfcUf 1147 GfAfcGfaGfgUfgGfcCfuGfuC 1148 cGfuCfdTsdTfgUfcdTsdT 116 116-134 acGfaCfaGfgCfcAfcCfuCf 1149CfGfaCfgAfgGfuGfgCfcUfgU 1150 gUfcGfdTsdT fcGfudTsdT 117 117-135cgAfcAfgGfcCfaCfcUfcGf 1151 CfCfgAfcGfaGfgUfgGfcCfuG 1152 uCfgGfdTsdTfuCfgdTsdT 118 118-136 gaCfaGfgCfcAfcCfuCfgUf 1153GfCfcGfaCfgAfgGfuGfgCfcU 1154 cGfgCfdTsdT fgUfcdTsdT 120 120-138caGfgCfcAfcCfuCfgUfcGf 1155 AfCfgCfcGfaCfgAfgGfuGfgC 1156 gCfgUfdTsdTfcUfgdTsdT 121 121-139 agGfcCfaCfcUfcGfuCfgGf 1157GfAfcGfcCfgAfcGfaGfgUfgG 1158 cGfuCfdTsdT fcCfudTsdT 122 122-140ggCfcAfcCfuCfgUfcGfgCf 1159 GfGfaCfgCfcGfaCfgAfgGfuG 1160 gUfcCfdTsdTfgCfcdTsdT 123 123-141 gcCfaCfcUfcGfuCfgGfcGf 1161CfGfgAfcGfcCfgAfcGfaGfgU 1162 uCfcGfdTsdT fgGfcdTsdT 124 124-142ccAfcCfuCfgUfcGfgCfgUf 1163 GfCfgGfaCfgCfcGfaCfgAfgG 1164 cCfgCfdTsdTfuGfgdTsdT 125 125-143 caCfcUfcGfuCfgGfcGfuCf 1165GfGfcGfgAfcGfcCfgAfcGfaG 1166 cGfcCfdTsdT fgUfgdTsdT 126 126-144acCfuCfgUfcGfgCfgUfcCf 1167 GfGfgCfgGfaCfgCfcGfaCfgA 1168 gCfcCfdTsdTfgGfudTsdT 127 127-145 ccUfcGfuCfgGfcGfuCfcGf 1169CfGfgGfcGfgAfcGfcCfgAfcG 1170 cCfcGfdTsdT faGfgdTsdT 128 128-146cuCfgUfcGfgCfgUfcCfgCf 1171 UfCfgGfgCfgGfaCfgCfcGfaC 1172 cCfgAfdTsdTfgAfgdTsdT 129 129-147 ucGfuCfgGfcGfuCfcGfcCf 1173CfUfcGfgGfcGfgAfcGfcCfgA 1174 cGfaGfdTsdT fcGfadTsdT 130 130-148cgUfcGfgCfgUfcCfgCfcCf 1175 AfCfuCfgGfgCfgGfaCfgCfcG 1176 gAfgUfdTsdTfaCfgdTsdT 131 131-149 guCfgGfcGfuCfcGfcCfcGf 1177GfAfcUfcGfgGfcGfgAfcGfcC 1178 aGfuCfdTsdT fgAfcdTsdT 132 132-150ucGfgCfgUfcCfgCfcCfgAf 1179 GfGfaCfuCfgGfgCfgGfaCfgC 1180 gUfcCfdTsdTfcGfadTsdT 135 135-153 gcGfuCfcGfcCfcGfaGfuCf 1181CfGfgGfgAfcUfcGfgGfcGfgA 1182 cCfcGfdTsdT fcGfcdTsdT 136 136-154cgUfcCfgCfcCfgAfgUfcCf 1183 GfCfgGfgGfaCfuCfgGfgCfgG 1184 cCfgCfdTsdTfaCfgdTsdT 141 141-159 gcCfcGfaGfuCfcCfcGfcCf 1185GfCfgAfgGfcGfgGfgAfcUfcG 1186 uCfgCfdTsdT fgGfcdTsdT 164 164-182aaCfgCfcAfcAfaCfcAfcCfg 1187 GfCfgCfgGfuGfgUfuGfuGfgC 1188 CfgCfdTsdTfgUfudTsdT 165 165-183 acGfcCfaCfaAfcCfaCfcGfc 1189UfGfcGfcGfgUfgGfuUfgUfg 1190 GfcAfdTsdT GfcGfudTsdT 166 166-184cgCfcAfcAfaCfcAfcCfgCf 1191 GfUfgCfgCfgGfuGfgUfuGfu 1192 gCfaCfdTsdTGfgCfgdTsdT 168 168-186 ccAfcAfaCfcAfcCfgCfgCfa 1193CfCfgUfgCfgCfgGfuGfgUfuG 1194 CfgGfdTsdT fuGfgdTsdT 169 169-187caCfaAfcCfaCfcGfcGfcAfc 1195 GfCfcGfuGfcGfcGfgUfgGfuU 1196 GfgCfdTsdTfgUfgdTsdT 170 170-188 acAfaCfcAfcCfgCfgCfaCfg 1197GfGfcCfgUfgCfgCfgGfuGfgU 1198 GfcCfdTsdT fuGfudTsdT 247 247-265auGfcGfaCfcCfuCfcGfgGf 1199 CfCfgUfcCfcGfgAfgGfgUfcG 1200 aCfgGfdTsdTfcAfudTsdT 248 248-266 ugCfgAfcCfcUfcCfgGfgAf 1201GfCfcGfuCfcCfgGfaGfgGfuC 1202 cGfgCfdTsdT fgCfadTsdT 249 249-267gcGfaCfcCfuCfcGfgGfaCf 1203 GfGfcCfgUfcCfcGfgAfgGfgU 1204 gGfcCfdTsdTfcGfcdTsdT 251 251-269 gaCfcCfuCfcGfgGfaCfgGf 1205CfCfgGfcCfgUfcCfcGfgAfgG 1206 cCfgGfdTsdT fgUfcdTsdT 252 252-270acCfcUfcCfgGfgAfcGfgCf 1207 CfCfcGfgCfcGfuCfcCfgGfaGf 1208 cGfgGfdTsdTgGfudTsdT 254 254-272 ccUfcCfgGfgAfcGfgCfcGf 1209GfCfcCfcGfgCfcGfuCfcCfgGf 1210 gGfgCfdTsdT aGfgdTsdT 329 329-347agAfaAfgUfuUfgCfcAfaGf 1211 GfUfgCfcUfuGfgCfaAfaCfuU 1212 gCfaCfdTsdTfuCfudTsdT 330 330-348 gaAfaGfuUfuGfcCfaAfgGf 1213CfGfuGfcCfuUfgGfcAfaAfcU 1214 cAfcGfdTsdT fuUfcdTsdT 332 332-350aaGfuUfuGfcCfaAfgGfcAf 1215 CfUfcGfuGfcCfuUfgGfcAfaA 1216 cGfaGfdTsdTfcUfudTsdT 333 333-351 agUfuUfgCfcAfaGfgCfaCf 1217AfCfuCfgUfgCfcUfuGfgCfaA 1218 gAfgUfdTsdT faCfudTsdT 334 334-352guUfuGfcCfaAfgGfcAfcGf 1219 UfAfcUfcGfuGfcCfuUfgGfcA 1220 aGfuAfdTsdTfaAfcdTsdT 335 335-353 uuUfgCfcAfaGfgCfaCfgAf 1221UfUfaCfuCfgUfgCfcUfuGfgC 1222 gUfaAfdTsdT faAfadTsdT 336 336-354uuGfcCfaAfgGfcAfcGfaGf 1223 GfUfuAfcUfcGfuGfcCfuUfgG 1224 uAfaCfdTsdTfcAfadTsdT 337 337-355 ugCfcAfaGfgCfaCfgAfgUf 1225UfGfuUfaCfuCfgUfgCfcUfuG 1226 aAfcAfdTsdT fgCfadTsdT 338 338-356gcCfaAfgGfcAfcGfaGfuAf 1227 UfUfgUfuAfcUfcGfuGfcCfuU 1228 aCfaAfdTsdTfgGfcdTsdT 361 361-379 acGfcAfgUfuGfgGfcAfcUf 1229CfAfaAfaGfuGfcCfcAfaCfuG 1230 uUfuGfdTsdT fcGfudTsdT 362 362-380cgCfaGfuUfgGfgCfaCfuUf 1231 UfCfaAfaAfgUfgCfcCfaAfcU 1232 uUfgAfdTsdTfgCfgdTsdT 363 363-381 gcAfgUfuGfgGfcAfcUfuUf 1233UfUfcAfaAfaGfuGfcCfcAfaC 1234 uGfaAfdTsdT fuGfcdTsdT 364 364-382caGfuUfgGfgCfaCfuUfuUf 1235 CfUfuCfaAfaAfgUfgCfcCfaA 1236 gAfaGfdTsdTfcUfgdTsdT 365 365-383 agUfuGfgGfcAfcUfuUfuGf 1237UfCfuUfcAfaAfaGfuGfcCfcA 1238 aAfgAfdTsdT faCfudTsdT 366 366-384guUfgGfgCfaCfuUfuUfgAf 1239 AfUfcUfuCfaAfaAfgUfgCfcC 1240 aGfaUfdTsdTfaAfcdTsdT 367 367-385 uuGfgGfcAfcUfuUfuGfaAf 1241GfAfuCfuUfcAfaAfaGfuGfcC 1242 gAfuCfdTsdT fcAfadTsdT 368 368-386ugGfgCfaCfuUfuUfgAfaGf 1243 UfGfaUfcUfuCfaAfaAfgUfgC 1244 aUfcAfdTsdTfcCfadTsdT 369 369-387 ggGfcAfcUfuUfuGfaAfgAf 1245AfUfgAfuCfuUfcAfaAfaGfuG 1246 uCfaUfdTsdT fcCfcdTsdT 377 377-395uuGfaAfgAfuCfaUfuUfuCf 1247 CfUfgAfgAfaAfaUfgAfuCfuU 1248 uCfaGfdTsdTfcAfadTsdT 379 379-397 gaAfgAfuCfaUfuUfuCfuCf 1249GfGfcUfgAfgAfaAfaUfgAfuC 1250 aGfcCfdTsdT fuUfcdTsdT 380 380-398aaGfaUfcAfuUfuUfcUfcAf 1251 AfGfgCfuGfaGfaAfaAfuGfaU 1252 gCfcUfdTsdTfcUfudTsdT 385 385-403 caUfuUfuCfuCfaGfcCfuCf 1253UfCfuGfgAfgGfcUfgAfgAfaA 1254 cAfgAfdTsdT faUfgdTsdT 394 394-412agCfcUfcCfaGfaGfgAfuGf 1255 UfGfaAfcAfuCfcUfcUfgGfaG 1256 uUfcAfdTsdTfgCfudTsdT 396 396-414 ccUfcCfaGfaGfgAfuGfuUf 1257AfUfuGfaAfcAfuCfcUfcUfgG 1258 cAfaUfdTsdT faGfgdTsdT 397 397-415cuCfcAfgAfgGfaUfgUfuCf 1259 UfAfuUfgAfaCfaUfcCfuCfuG 1260 aAfuAfdTsdTfgAfgdTsdT 401 401-419 agAfgGfaUfgUfuCfaAfuAf 1261CfAfgUfuAfuUfgAfaCfaUfcC 1262 aCfuGfdTsdT fuCfudTsdT 403 403-421agGfaUfgUfuCfaAfuAfaCf 1263 CfAfcAfgUfuAfuUfgAfaCfaU 1264 uGfuGfdTsdTfcCfudTsdT 407 407-425 ugUfuCfaAfuAfaCfuGfuGf 1265AfCfcUfcAfcAfgUfuAfuUfgA 1266 aGfgUfdTsdT faCfadTsdT 409 409-427uuCfaAfuAfaCfuGfuGfaGf 1267 CfCfaCfcUfcAfcAfgUfuAfuU 1268 gUfgGfdTsdTfgAfadTsdT 410 410-428 ucAfaUfaAfcUfgUfgAfgGf 1269AfCfcAfcCfuCfaCfaGfuUfaUf 1270 uGfgUfdTsdT uGfadTsdT 411 411-429caAfuAfaCfuGfuGfaGfgUf 1271 GfAfcCfaCfcUfcAfcAfgUfuA 1272 gGfuCfdTsdTfuUfgdTsdT 412 412-430 aaUfaAfcUfgUfgAfgGfuGf 1273GfGfaCfcAfcCfuCfaCfaGfuUf 1274 gUfcCfdTsdT aUfudTsdT 413 413-431auAfaCfuGfuGfaGfgUfgGf 1275 AfGfgAfcCfaCfcUfcAfcAfgU 1276 uCfcUfdTsdTfuAfudTsdT 414 414-432 uaAfcUfgUfgAfgGfuGfgUf 1277AfAfgGfaCfcAfcCfuCfaCfaGf 1278 cCfuUfdTsdT uUfadTsdT 416 416-434acUfgUfgAfgGfuGfgUfcCf 1279 CfCfaAfgGfaCfcAfcCfuCfaCf 1280 uUfgGfdTsdTaGfudTsdT 418 418-436 ugUfgAfgGfuGfgUfcCfuUf 1281UfCfcCfaAfgGfaCfcAfcCfuCf 1282 gGfgAfdTsdT aCfadTsdT 419 419-437guGfaGfgUfgGfuCfcUfuGf 1283 UfUfcCfcAfaGfgAfcCfaCfcUf 1284 gGfaAfdTsdTcAfcdTsdT 425 425-443 ugGfuCfcUfuGfgGfaAfuUf 1285UfCfcAfaAfuUfcCfcAfaGfgA 1286 uGfgAfdTsdT fcCfadTsdT 431 431-449uuGfgGfaAfuUfuGfgAfaAf 1287 GfUfaAfuUfuCfcAfaAfuUfcC 1288 uUfaCfdTsdTfcAfadTsdT 432 432-450 ugGfgAfaUfuUfgGfaAfaUf 1289GfGfuAfaUfuUfcCfaAfaUfuC 1290 uAfcCfdTsdT fcCfadTsdT 433 433-451ggGfaAfuUfuGfgAfaAfuUf 1291 AfGfgUfaAfuUfuCfcAfaAfuU 1292 aCfcUfdTsdTfcCfcdTsdT 434 434-452 ggAfaUfuUfgGfaAfaUfuAf 1293UfAfgGfuAfaUfuUfcCfaAfaU 1294 cCfuAfdTsdT fuCfcdTsdT 458 458-476agAfgGfaAfuUfaUfgAfuCf 1295 GfAfaAfgAfuCfaUfaAfuUfcC 1296 uUfuCfdTsdTfuCfudTsdT 459 459-477 gaGfgAfaUfuAfuGfaUfcUf 1297GfGfaAfaGfaUfcAfuAfaUfuC 1298 uUfcCfdTsdT fcUfcdTsdT 463 463-481aaUfuAfuGfaUfcUfuUfcCf 1299 AfGfaAfgGfaAfaGfaUfcAfuA 1300 uUfcUfdTsdTfaUfudTsdT 464 464-482 auUfaUfgAfuCfuUfuCfcUf 1301AfAfgAfaGfgAfaAfgAfuCfaU 1302 uCfuUfdTsdT faAfudTsdT 466 466-484uaUfgAfuCfuUfuCfcUfuCf 1303 UfUfaAfgAfaGfgAfaAfgAfuC 1304 uUfaAfdTsdTfaUfadTsdT 468 468-486 ugAfuCfuUfuCfcUfuCfuUf 1305CfUfuUfaAfgAfaGfgAfaAfgA 1306 aAfaGfdTsdT fuCfadTsdT 471 471-489ucUfuUfcCfuUfcUfuAfaAf 1307 GfGfuCfuUfuAfaGfaAfgGfaA 1308 gAfcCfdTsdTfaGfadTsdT 476 476-494 ccUfuCfuUfaAfaGfaCfcAf 1309UfGfgAfuGfgUfcUfuUfaAfg 1310 uCfcAfdTsdT AfaGfgdTsdT 477 477-495cuUfcUfuAfaAfgAfcCfaUf 1311 CfUfgGfaUfgGfuCfuUfuAfaG 1312 cCfaGfdTsdTfaAfgdTsdT 479 479-497 ucUfuAfaAfgAfcCfaUfcCf 1313UfCfcUfgGfaUfgGfuCfuUfuA 1314 aGfgAfdTsdT faGfadTsdT 481 481-499uuAfaAfgAfcCfaUfcCfaGf 1315 CfCfuCfcUfgGfaUfgGfuCfuU 1316 gAfgGfdTsdTfuAfadTsdT 482 482-500 uaAfaGfaCfcAfuCfcAfgGf 1317AfCfcUfcCfuGfgAfuGfgUfcU 1318 aGfgUfdTsdT fuUfadTsdT 492 492-510ccAfgGfaGfgUfgGfcUfgGf 1319 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1745UfCfgCfaCfuUfcUfuAfcAfcU 1746 gCfgAfdTsdT fuGfcdTsdT 1219 1219-1237aaGfuGfuAfaGfaAfgUfgCf 1747 CfUfuCfgCfaCfuUfcUfuAfcA 1748 gAfaGfdTsdTfcUfudTsdT 1220 1220-1238 agUfgUfaAfgAfaGfuGfcGf 1749CfCfuUfcGfcAfcUfuCfuUfaC 1750 aAfgGfdTsdT faCfudTsdT 1221 1221-1239guGfuAfaGfaAfgUfgCfgAf 1751 CfCfcUfuCfgCfaCfuUfcUfuA 1752 aGfgGfdTsdTfcAfcdTsdT 1222 1222-1240 ugUfaAfgAfaGfuGfcGfaAf 1753GfCfcCfuUfcGfcAfcUfuCfuU 1754 gGfgCfdTsdT faCfadTsdT 1223 1223-1241guAfaGfaAfgUfgCfgAfaGf 1755 GfGfcCfcUfuCfgCfaCfuUfcU 1756 gGfcCfdTsdTfuAfcdTsdT 1224 1224-1242 uaAfgAfaGfuGfcGfaAfgGf 1757AfGfgCfcCfuUfcGfcAfcUfuC 1758 gCfcUfdTsdT fuUfadTsdT 1225 1225-1243aaGfaAfgUfgCfgAfaGfgGf 1759 AfAfgGfcCfcUfuCfgCfaCfuU 1760 cCfuUfdTsdTfcUfudTsdT 1226 1226-1244 agAfaGfuGfcGfaAfgGfgCf 1761CfAfaGfgCfcCfuUfcGfcAfcU 1762 cUfuGfdTsdT fuCfudTsdT 1229 1229-1247agUfgCfgAfaGfgGfcCfuUf 1763 CfGfgCfaAfgGfcCfcUfuCfgC 1764 gCfcGfdTsdTfaCfudTsdT 1230 1230-1248 guGfcGfaAfgGfgCfcUfuGf 1765GfCfgGfcAfaGfgCfcCfuUfcG 1766 cCfgCfdTsdT fcAfcdTsdT 1231 1231-1249ugCfgAfaGfgGfcCfuUfgCf 1767 UfGfcGfgCfaAfgGfcCfcUfuC 1768 cGfcAfdTsdTfgCfadTsdT 1232 1232-1250 gcGfaAfgGfgCfcUfuGfcCf 1769UfUfgCfgGfcAfaGfgCfcCfuU 1770 gCfaAfdTsdT fcGfcdTsdT 1233 1233-1251cgAfaGfgGfcCfuUfgCfcGf 1771 UfUfuGfcGfgCfaAfgGfcCfcU 1772 cAfaAfdTsdTfuCfgdTsdT 1235 1235-1253 aaGfgGfcCfuUfgCfcGfcAf 1773AfCfuUfuGfcGfgCfaAfgGfcC 1774 aAfgUfdTsdT fcUfudTsdT 1236 1236-1254agGfgCfcUfuGfcCfgCfaAf 1775 CfAfcUfuUfgCfgGfcAfaGfgC 1776 aGfuGfdTsdTfcCfudTsdT 1237 1237-1255 ggGfcCfuUfgCfcGfcAfaAf 1777AfCfaCfuUfuGfcGfgCfaAfgG 1778 gUfgUfdTsdT fcCfcdTsdT 1238 1238-1256ggCfcUfuGfcCfgCfaAfaGf 1779 CfAfcAfcUfuUfgCfgGfcAfaG 1780 uGfuGfdTsdTfgCfcdTsdT 1239 1239-1257 gcCfuUfgCfcGfcAfaAfgUf 1781AfCfaCfaCfuUfuGfcGfgCfaA 1782 gUfgUfdTsdT fgGfcdTsdT 1241 1241-1259cuUfgCfcGfcAfaAfgUfgUf 1783 UfUfaCfaCfaCfuUfuGfcGfgC 1784 gUfaAfdTsdTfaAfgdTsdT 1261 1261-1279 ggAfaUfaGfgUfaUfuGfgUf 1785AfUfuCfaCfcAfaUfaCfcUfaUf 1786 gAfaUfdTsdT uCfcdTsdT 1262 1262-1280gaAfuAfgGfuAfuUfgGfuG 1787 AfAfuUfcAfcCfaAfuAfcCfuA 1788 faAfuUfdTsdTfuUfcdTsdT 1263 1263-1281 aaUfaGfgUfaUfuGfgUfgAf 1789AfAfaUfuCfaCfcAfaUfaCfcUf 1790 aUfuUfdTsdT aUfudTsdT 1264 1264-1282auAfgGfuAfuUfgGfuGfaAf 1791 UfAfaAfuUfcAfcCfaAfuAfcC 1792 uUfuAfdTsdTfuAfudTsdT 1266 1266-1284 agGfuAfuUfgGfuGfaAfuUf 1793UfUfuAfaAfuUfcAfcCfaAfuA 1794 uAfaAfdTsdT fcCfudTsdT 1267 1267-1285ggUfaUfuGfgUfgAfaUfuUf 1795 CfUfuUfaAfaUfuCfaCfcAfaU 1796 aAfaGfdTsdTfaCfcdTsdT 1289 1289-1307 caCfuCfuCfcAfuAfaAfuGf 1797GfUfaGfcAfuUfuAfuGfgAfg 1798 cUfaCfdTsdT AfgUfgdTsdT 1313 1313-1331uuAfaAfcAfcUfuCfaAfaAf 1799 CfAfgUfuUfuUfgAfaGfuGfu 1800 aCfuGfdTsdTUfuAfadTsdT 1320 1320-1338 cuUfcAfaAfaAfcUfgCfaCf 1801GfGfaGfgUfgCfaGfuUfuUfuG 1802 cUfcCfdTsdT faAfgdTsdT 1321 1321-1339uuCfaAfaAfaCfuGfcAfcCf 1803 UfGfgAfgGfuGfcAfgUfuUfu 1804 uCfcAfdTsdTUfgAfadTsdT 1322 1322-1340 ucAfaAfaAfcUfgCfaCfcUf 1805AfUfgGfaGfgUfgCfaGfuUfuU 1806 cCfaUfdTsdT fuGfadTsdT 1323 1323-1341caAfaAfaCfuGfcAfcCfuCf 1807 GfAfuGfgAfgGfuGfcAfgUfu 1808 cAfuCfdTsdTUfuUfgdTsdT 1324 1324-1342 aaAfaAfcUfgCfaCfcUfcCfa 1809UfGfaUfgGfaGfgUfgCfaGfuU 1810 UfcAfdTsdT fuUfudTsdT 1328 1328-1346acUfgCfaCfcUfcCfaUfcAf 1811 CfCfaCfuGfaUfgGfaGfgUfgC 1812 gUfgGfdTsdTfaGfudTsdT 1332 1332-1350 caCfcUfcCfaUfcAfgUfgGf 1813AfUfcGfcCfaCfuGfaUfgGfaG 1814 cGfaUfdTsdT fgUfgdTsdT 1333 1333-1351acCfuCfcAfuCfaGfuGfgCf 1815 GfAfuCfgCfcAfcUfgAfuGfgA 1816 gAfuCfdTsdTfgGfudTsdT 1335 1335-1353 cuCfcAfuCfaGfuGfgCfgAf 1817GfAfgAfuCfgCfcAfcUfgAfuG 1818 uCfuCfdTsdT fgAfgdTsdT 1338 1338-1356caUfcAfgUfgGfcGfaUfcUf 1819 GfUfgGfaGfaUfcGfcCfaCfuG 1820 cCfaCfdTsdTfaUfgdTsdT 1344 1344-1362 ugGfcGfaUfcUfcCfaCfaUf 1821CfAfgGfaUfgUfgGfaGfaUfcG 1822 cCfuGfdTsdT fcCfadTsdT 1345 1345-1363ggCfgAfuCfuCfcAfcAfuCf 1823 GfCfaGfgAfuGfuGfgAfgAfuC 1824 cUfgCfdTsdTfgCfcdTsdT 1346 1346-1364 gcGfaUfcUfcCfaCfaUfcCf 1825GfGfcAfgGfaUfgUfgGfaGfaU 1826 uGfcCfdTsdT fcGfcdTsdT 1347 1347-1365cgAfuCfuCfcAfcAfuCfcUf 1827 CfGfgCfaGfgAfuGfuGfgAfgA 1828 gCfcGfdTsdTfuCfgdTsdT 1348 1348-1366 gaUfcUfcCfaCfaUfcCfuGf 1829CfCfgGfcAfgGfaUfgUfgGfaG 1830 cCfgGfdTsdT faUfcdTsdT 1353 1353-1371ccAfcAfuCfcUfgCfcGfgUf 1831 UfGfcCfaCfcGfgCfaGfgAfuG 1832 gGfcAfdTsdTfuGfgdTsdT 1354 1354-1372 caCfaUfcCfuGfcCfgGfuGf 1833AfUfgCfcAfcCfgGfcAfgGfaU 1834 gCfaUfdTsdT fgUfgdTsdT 1355 1355-1373acAfuCfcUfgCfcGfgUfgGf 1835 AfAfuGfcCfaCfcGfgCfaGfgA 1836 cAfuUfdTsdTfuGfudTsdT 1357 1357-1375 auCfcUfgCfcGfgUfgGfcAf 1837UfAfaAfuGfcCfaCfcGfgCfaG 1838 uUfuAfdTsdT fgAfudTsdT 1360 1360-1378cuGfcCfgGfuGfgCfaUfuUf 1839 CfCfcUfaAfaUfgCfcAfcCfgGf 1840 aGfgGfdTsdTcAfgdTsdT 1361 1361-1379 ugCfcGfgUfgGfcAfuUfuAf 1841CfCfcCfuAfaAfuGfcCfaCfcGf 1842 gGfgGfdTsdT gCfadTsdT 1362 1362-1380gcCfgGfuGfgCfaUfuUfaGf 1843 AfCfcCfcUfaAfaUfgCfcAfcCf 1844 gGfgUfdTsdTgGfcdTsdT 1363 1363-1381 ccGfgUfgGfcAfuUfuAfgGf 1845CfAfcCfcCfuAfaAfuGfcCfaCf 1846 gGfuGfdTsdT cGfgdTsdT 1366 1366-1384guGfgCfaUfuUfaGfgGfgUf 1847 AfGfuCfaCfcCfcUfaAfaUfgCf 1848 gAfcUfdTsdTcAfcdTsdT 1369 1369-1387 gcAfuUfuAfgGfgGfuGfaCf 1849AfGfgAfgUfcAfcCfcCfuAfaA 1850 uCfcUfdTsdT fuGfcdTsdT 1370 1370-1388caUfuUfaGfgGfgUfgAfcUf 1851 AfAfgGfaGfuCfaCfcCfcUfaA 1852 cCfuUfdTsdTfaUfgdTsdT 1371 1371-1389 auUfuAfgGfgGfuGfaCfuCf 1853GfAfaGfgAfgUfcAfcCfcCfuA 1854 cUfuCfdTsdT faAfudTsdT 1372 1372-1390uuUfaGfgGfgUfgAfcUfcCf 1855 UfGfaAfgGfaGfuCfaCfcCfcU 1856 uUfcAfdTsdTfaAfadTsdT 1373 1373-1391 uuAfgGfgGfuGfaCfuCfcUf 1857GfUfgAfaGfgAfgUfcAfcCfcC 1858 uCfaCfdTsdT fuAfadTsdT 1374 1374-1392uaGfgGfgUfgAfcUfcCfuUf 1859 UfGfuGfaAfgGfaGfuCfaCfcC 1860 cAfcAfdTsdTfcUfadTsdT 1404 1404-1422 ucUfgGfaUfcCfaCfaGfgAf 1861CfAfgUfuCfcUfgUfgGfaUfcC 1862 aCfuGfdTsdT faGfadTsdT 1408 1408-1426gaUfcCfaCfaGfgAfaCfuGf 1863 UfAfuCfcAfgUfuCfcUfgUfgG 1864 gAfuAfdTsdTfaUfcdTsdT 1409 1409-1427 auCfcAfcAfgGfaAfcUfgGf 1865AfUfaUfcCfaGfuUfcCfuGfuG 1866 aUfaUfdTsdT fgAfudTsdT 1411 1411-1429ccAfcAfgGfaAfcUfgGfaUf 1867 GfAfaUfaUfcCfaGfuUfcCfuG 1868 aUfuCfdTsdTfuGfgdTsdT 1412 1412-1430 caCfaGfgAfaCfuGfgAfuAf 1869AfGfaAfuAfuCfcAfgUfuCfcU 1870 uUfcUfdTsdT fgUfgdTsdT 1419 1419-1437acUfgGfaUfaUfuCfuGfaAf 1871 GfGfuUfuUfcAfgAfaUfaUfcC 1872 aAfcCfdTsdTfaGfudTsdT 1426 1426-1444 auUfcUfgAfaAfaCfcGfuAf 1873CfCfuUfuAfcGfgUfuUfuCfaG 1874 aAfgGfdTsdT faAfudTsdT 1427 1427-1445uuCfuGfaAfaAfcCfgUfaAf 1875 UfCfcUfuUfaCfgGfuUfuUfcA 1876 aGfgAfdTsdTfgAfadTsdT 1430 1430-1448 ugAfaAfaCfcGfuAfaAfgGf 1877AfUfuUfcCfuUfuAfcGfgUfuU 1878 aAfaUfdTsdT fuCfadTsdT 1431 1431-1449gaAfaAfcCfgUfaAfaGfgAf 1879 GfAfuUfuCfcUfuUfaCfgGfuU 1880 aAfuCfdTsdTfuUfcdTsdT siRNA Sequence with Chemical Modification Info lower case (n)= 2′-O—Me; Nf = 2′-F; dT = deoxy-T residue; s = phosphorothioatebackbone modification; iB = inverted abasic

Example 2. Evaluation of In Vitro Potency of Anti-EGFR siRNAs

Each of the anti-EGFR siRNAs in Table 4 were transfected in each ofthree human non-small cell lung cancer (NSCLC) cell lines with theindicated EGFR mutational status:

-   -   H358: wild-type    -   H1650: Exon19 ΔE746-A750 deletion    -   H1975: Exon21 L858R and Exon20 T790M

At a single final concentration of 5 nM, each siRNA was formulated witha commercially-available transfection reagent (Lipofectamine RNAiMAX,Life Technologies) according to the manufacturer's “forwardtransfection” instructions. Cells were plated 24 h prior to transfectionin duplicate within 24-well tissue culture plates. At 24 h (H1650 andH1975) or 48 h (H358) post-transfection, RNA was harvested from cells inall wells using a Qiagen RNeasy® Plus Mini Kit or Stratec InviTrap® RNACell HTS96 kit. The concentration of each isolated RNA was determinedvia A260 measurement using a NanoDrop spectrophotometer. RNA sampleswere then reverse transcribed to cDNA using the High Capacity RNA tocDNA Kit (Life Technologies) according to the manufacturer'sinstructions. cDNA samples were then evaluated by qPCR usingEGFR-specific probes with results normalized to endogenous (3-actin andquantified using the standard 2^(−ΔΔCt) method. EGFR mRNA levelsnormalized to expression in controls were determined.

TABLE 4 qPCR, qPCR, qPCR, H358, H1650, H1975, 5 nM 5 nM 5 nM SEQ SEQ %Rel % Rel % Rel Avidity sense strand ID antisense strand ID EGFR EGFREGFR ID# sequence (5′-3′) NO: sequence (5′-3′) NO: mRNA mRNA mRNA R-1006agUfuUfgCfcAfaG 1217 AfCfuCfgUfgCfcUfu 1218 7.80% 5.50% 12.20%fgCfaCfgAfgUfdTs GfgCfaAfaCfudTsdT dT R-1010 agCfaGfuCfuUfaUf 1373AfUfaGfuUfaGfaUfa 1374 3.80% 3.50% 8.60% cUfaAfcUfaUfdTsdTAfgAfcUfgCfudTsdT R-1018 gcCfgUfgCfgGfuU 1447 UfGfuUfgCfuGfaAfc 14487.40% 6.50% 13.00% fcAfgCfaAfcAfdTs CfgCfaCfgGfcdTsdT dT R-1011gaUfgCfaAfaUfaAf 1395 GfUfcCfgGfuUfuUfa 1396 9.70% 4.80% 10.10%aAfcCfgGfaCfdTsdT UfuUfgCfaUfcdTsdT R-1001 gaCfgAfcAfgGfcCf 1147GfAfcGfaGfgUfgGfc 1148 95.40% 118.90% 109.30% aCfcUfcGfuCfdTsdTCfuGfuCfgUfcdTsdT R-1002 acGfaCfaGfgCfcAf 1149 CfGfaCfgAfgGfuGfg 115069.30% 98.10% 112.20% cCfuCfgUfcGfdTsdT CfcUfgUfcGfudTsdT R-1003ccAfcCfuCfgUfcGf 1163 GfCfgGfaCfgCfcGfa 1164 59.10% 81.50% 92.40%gCfgUfcCfgCfdTsdT CfgAfgGfuGfgdTsdT R-1004 guCfgGfcGfuCfcG 1177GfAfcUfcGfgGfcGfg 1178 93.40% 121.30% 123.40% fcCfcGfaGfuCfdTsAfcGfcCfgAfcdTsdT dT R-1005 ccAfcAfaCfcAfcCf 1193 CfCfgUfgCfgCfgGfu 119470.30% 90.80% 124.10% gCfgCfaCfgGfdTsdT GfgUfuGfuGfgdTsdT R-1007uuUfgCfcAfaGfgC 1221 UfUfaCfuCfgUfgCfc 1222 73.40% 75.70% 107.10%faCfgAfgUfaAfdTs UfuGfgCfaAfadTsdT dT R-1008 auGfcCfuUfaGfcAf 1359GfAfuAfaGfaCfuGfc 1360 8.50% 7.70% 30.60% gUfcUfuAfuCfdTsUfaAfgGfcAfudTsdT dT R-1009 uuAfgCfaGfuCfuU 1369 AfGfuUfaGfaUfaAfg 137010.50% 10.60% 25.70% faUfcUfaAfcUfdTs AfcUfgCfuAfadTsdT dT R-1012auGfcAfaAfuAfaA 1397 AfGfuCfcGfgUfuUfu 1398 22.30% 20.40% 41.30%faCfcGfgAfcUfdTs AfuUfuGfcAfudTsdT dT R-1013 ugCfaAfaUfaAfaAf 1399CfAfgUfcCfgGfuUfu 1400 22.80% 24.80% 50.00% cCfgGfaCfuGfdTsdTUfaUfuUfgCfadTsdT R-1014 agGfaAfaUfcCfuGf 1419 GfCfgCfcAfuGfcAfg 142017.50% 12.10% 24.30% cAfuGfgCfgCfdTs GfaUfuUfcCfudTsdT dT R-1015ggAfaAfuCfcUfgC 1421 GfGfcGfcCfaUfgCfa 1422 41.30% 41.10% 66.00%faUfgGfcGfcCfdTs GfgAfuUfuCfcdTsdT dT R-1016 aaAfuCfcUfgCfaUf 1425AfCfgGfcGfcCfaUfg 1426 89.90% 89.70% 94.40% gGfcGfcCfgUfdTsCfaGfgAfuUfudTsdT dT R-1017 ugCfaUfgGfcGfcCf 1433 AfAfcCfgCfaCfgGfc 143481.40% 99.20% 108.60% gUfgCfgGfuUfdTs GfcCfaUfgCfadTsdT dT R-1019gcGfgUfuCfaGfcA 1453 AfGfgGfuUfgUfuGfc 1454 30.20% 24.70% 45.10%faCfaAfcCfcUfdTs UfgAfaCfcGfcdTsdT dT R-1020 agUfgGfcGfgGfaC 1475CfUfgAfcUfaUfgUfc 1476 71.80% 60.50% 82.50% faUfaGfuCfaGfdTsCfcGfcCfaCfudTsdT dT R-1021 cuCfaGfcAfaCfaUf 1481 CfCfaUfcGfaCfaUfg 148261.70% 99.40% 91.10% gUfcGfaUfgGfdTs UfuGfcUfgAfgdTsdT dT R-1022gaUfcCfaAfgCfuGf 1517 CfAfuUfgGfgAfcAfg 1518 26.40% 24.50% 58.60%uCfcCfaAfuGfdTsdT CfuUfgGfaUfcdTsdT R-1023 ccAfaGfcUfgUfcCf 1523UfCfcCfaUfuGfgGfa 1524 61.30% 84.80% 99.00% cAfaUfgGfgAfdTsCfaGfcUfuGfgdTsdT dT R-1024 gcCfuGfgUfcUfgC 1565 AfAfuUfuGfcGfgCfa 156620.00% 15.30% 23.60% fcGfcAfaAfuUfdTs GfaCfcAfgGfcdTsdT dT R-1025ccUfgGfuCfuGfcCf 1567 GfAfaUfuUfgCfgGfc 1568 81.40% 77.40% 95.00%gCfaAfaUfuCfdTsdT AfgAfcCfaGfgdTsdT R-1026 cuGfgUfcUfgCfcG 1569GfGfaAfuUfuGfcGfg 1570 20.40% 24.20% 65.90% fcAfaAfuUfcCfdTsCfaGfaCfcAfgdTsdT dT R-1027 ugGfuCfuGfcCfgC 1571 CfGfgAfaUfuUfgCfg 157215.40% 11.30% 41.20% faAfaUfuCfcGfdTs GfcAfgAfcCfadTsdT dT R-1028cuGfcCfgCfaAfaUf 1577 GfUfcUfcGfgAfaUfu 1578 39.00% 39.70% 15.60%uCfcGfaGfaCfdTsdT UfgCfgGfcAfgdTsdT R-1029 ugCfcGfcAfaAfuU 1579CfGfuCfuCfgGfaAfu 1580 28.70% 18.90% 30.90% fcCfgAfgAfcGfdTsUfuGfcGfgCfadTsdT dT R-1030 ccCfcAfcUfcAfuGf 1603 UfUfgUfaGfaGfcAfu 160434.00% 34.30% 17.10% cUfcUfaCfaAfdTsdT GfaGfuGfgGfgdTsdT R-1031agAfuGfgAfuGfuG 1613 UfCfgGfgGfuUfcAfc 1614 17.80% 11.20% 52.60%faAfcCfcCfgAfdTs AfuCfcAfuCfudTsdT dT R-1032 cuGfcGfuGfaAfgA 1647GfGfgAfcAfcUfuCfu 1648 30.50% 13.40% 40.40% faGfuGfuCfcCfdTsUfcAfcGfcAfgdTsdT dT R-1033 gaAfgAfaGfuGfuC 1659 AfUfuAfcGfgGfgAfc 166011.10% 5.90% 46.80% fcCfcGfuAfaUfdTs AfcUfuCfuUfcdTsdT dT R-1034auGfuGfgUfgAfcA 1691 CfCfgUfgAfuCfuGfu 1692 27.90% 34.70% 9.00%fgAfuCfaCfgGfdTs CfaCfcAfcAfudTsdT dT R-1035 ugGfuGfaCfaGfaU 1693GfAfgCfcGfuGfaUfc 1694 59.00% 48.80% 82.20% fcAfcGfgCfuCfdTsUfgUfcAfcCfadTsdT dT R-1036 guGfaCfaGfaUfcAf 1695 AfCfgAfgCfcGfuGfa 169621.40% 27.10% 91.20% cGfgCfuCfgUfdTs UfcUfgUfcAfcdTsdT dT R-1037ugAfcAfgAfuCfaC 1697 CfAfcGfaGfcCfgUfg 1698 99.70% 90.20% 94.40%fgGfcUfcGfuGfdTs AfuCfuGfuCfadTsdT dT R-1038 gaCfaGfaUfcAfcGf 1699GfCfaCfgAfgCfcGfu 1700 25.60% 10.90% 88.30% gCfuCfgUfgCfdTsGfaUfcUfgUfcdTsdT dT R-1039 acAfgAfuCfaCfgGf 1701 CfGfcAfcGfaGfcCfg 170294.80% 98.50% 95.40% cUfcGfuGfcGfdTs UfgAfuCfuGfudTsdT dT R-1040caGfaUfcAfcGfgCf 1703 AfCfgCfaCfgAfgCfc 1704 108.70% 98.10% 90.00%uCfgUfgCfgUfdTs GfuGfaUfcUfgdTsdT dT R-1041 ucAfcGfgCfuCfgU 1711UfCfgGfaCfgCfaCfg 1712 93.70% 76.20% 38.30% fgCfgUfcCfgAfdTsAfgCfcGfuGfadTsdT dT R-1042 caCfgGfcUfcGfuGf 1713 CfUfcGfgAfcGfcAfc 171486.90% 92.30% 92.70% cGfuCfcGfaGfdTsdT GfaGfcCfgUfgdTsdT R-1043gaCfgGfcGfuCfcGf 1729 UfAfcAfcUfuGfcGfg 1730 60.00% 61.50% 8.00%cAfaGfuGfuAfdTs AfcGfcCfgUfcdTsdT dT R-1044 acGfgCfgUfcCfgCf 1731UfUfaCfaCfuUfgCfg 1732 21.00% 36.60% 90.90% aAfgUfgUfaAfdTsGfaCfgCfcGfudTsdT dT R-1045 gcGfuCfcGfcAfaGf 1733 UfUfcUfuAfcAfcUfu 173432.20% 21.50% 101.60% uGfuAfaGfaAfdTs GfcGfgAfcGfcdTsdT dT R-1046aaGfuGfuAfaGfaA 1747 CfUfuCfgCfaCfuUfc 1748 15.80% 10.10% 34.90%fgUfgCfgAfaGfdTs UfuAfcAfcUfudTsdT dT R-1047 cgAfaGfgGfcCfuU 1771UfUfuGfcGfgCfaAfg 1772 15.10% 10.00% 24.90% fgCfcGfcAfaAfdTsGfcCfcUfuCfgdTsdT dT R-1048 aaGfgGfcCfuUfgCf 1773 AfCfuUfuGfcGfgCfa 1774105.00% 96.80% 86.90% cGfcAfaAfgUfdTs AfgGfcCfcUfudTsdT dT R-1049acCfuCfcAfuCfaGf 1815 GfAfuCfgCfcAfcUfg 1816 89.50% 97.60% 102.90%uGfgCfgAfuCfdTs AfuGfgAfgGfudTsdT dT R-1050 cuCfcAfuCfaGfuGf 1817GfAfgAfuCfgCfcAfc 1818 39.00% 24.80% 68.60% gCfgAfuCfuCfdTsUfgAfuGfgAfgdTsdT dT R-1051 gcCfgGfuGfgCfaU 1843 AfCfcCfcUfaAfaUfg 184431.90% 30.40% 81.30% fuUfaGfgGfgUfdTs CfcAfcCfgGfcdTsdT dT

A list of the relative normalized EGFR mRNA levels, expressed as apercentage of control cells, for the initial single-concentrationtesting in all three cell lines is presented in Table 4. Of the siRNAcandidates tested, four (4) achieved ≧80% down-regulation in all threecell lines tested. Based upon results obtained from this initialsingle-concentration experiment, we selected a total of four (4)candidate siRNAs (hs ID #333, #603, #622, and #685) formulti-concentration testing, which allowed determination of IC₅₀ (i.e.,the concentration required to reduce EGFR expression by 50%), in each ofthese three cell lines. Cells were transfected as above atconcentrations starting from 5000 pM using serial dilutions. 48 hourspost-transfection, RNA was harvested and normalized EGFR mRNA levelswere quantified as described above. Results are presented in Table 5,and all four siRNAs had comparable potency in term of silencing the mRNAtarget.

TABLE 5 qPCR, qPCR, qPCR, Viability Viability, Avidity H358, H1650,H1975, H358, H1975, ID# IC50, pM IC50, pM IC50, pM IC50, pM IC50, pMR-1006 21 3.5 181 395 21 R-1010 17 7 131 R-1018 21 10.4 99 R-1011 3010.6 103 195 24

hs ID #333 and #622 were evaluated to see if reduction in EGFR mRNAresulted in reduced cell viability. Knocking down EGFR mRNA is known toaffect cell viability and proliferation, particularly in cell linescarrying EGFR activating mutations, such as H1975. 3000 H358 and H1975cells were plated in 384-well plates and increasing concentrations ofsiRNA-#333 and #622 in the presence with RNAiMAX were added to the cellculture. At 4 days post transfection, new media and siRNAs withtransfection reagent were added to the cell culture plates. After atotal of seven days (3 days post 2^(nd) transfection), cell viabilitywas assessed with CellTiter-Blue® (Promega), according to themanufacturer's instructions and is reported in table 2. Knocking downEGFR mRNA, as shown in table 2, had a dramatic effect on cell viabilityon both cell lines.

Next, an array of chemical modification patterns were introduced tosiRNA-#333 (Table 6) and their effect on EGFR mRNA were tested in H358and H1975 cells after transfection with RNAiMAX as described above.Modification patterns used in R-1068 and R-1105 caused significantactivity loss while the others showed comparable IC₅₀ values to theoriginal siRNA-#333 in both cell lines tested.

TABLE 6 qPCR, qPCR, SEQ SEQ H358, H1650, Avidity sense strand sequenceID antisense strand ID IC50, IC50, ID (5′-3′) NO: sequence (5′-3′) NO:pM pM R-1067 AGUUUGCCAAGGCA 1881 ACUCGUGCCUUG 1882 12.7 167.7 CGAGUdTsdTGCAAACUdTsdT R-1068 AaguuuGccAAGGcacGA 1883 ACUCGUGCCUUG 1884304.8 >1000 GudTsdT GcAAACUdTsdT R-1102 iBagUfuUfgCfcAfaGfgC 1885AfCfuCfgUfgCfcUfu 1886 18.7 95.3 faCfgAfgUfdTsdTiB GfgCfaAfaCfudTsdTR-1103 iBagUfuUfgCfcAfaGfgC 1887 AfsCfsusCfgUfgCfc 1888 48.4 99.1faCfgAfgUfdTsdTiB UfuGfgCfaAfaCfudT sdT R-1104 iBagUfuUfgCfcAfaGfgC 1889aCfuCfgUfgCfcUfuG 1890 60.5 145.9 faCfgAfgUfdTsdTiB fgCfaAfaCfudTsdTR-1105 iBagUfuUfgCfcAfaGfgC 1891 asCfsusCfgUfgCfcUf 1892 167.4 546.7faCfgAfgUfdTsdTiB uGfgCfaAfaCfudTsdT

While preferred embodiments of the present disclosure have been shownand described herein, it will be obvious to those skilled in the artthat such embodiments are provided by way of example only. Numerousvariations, changes, and substitutions will now occur to those skilledin the art without departing from the disclosure. It should beunderstood that various alternatives to the embodiments of thedisclosure described herein may be employed in practicing thedisclosure. It is intended that the following claims define the scope ofthe disclosure and that methods and structures within the scope of theseclaims and their equivalents be covered thereby.

What is claimed is:
 1. A polynucleic acid molecule that mediates RNAinterference against EGFR, wherein the polynucleic acid moleculehybridizes to a EGFR target sequence selected from SEQ ID NOs: 1-376with less than 4 mismatched bases, wherein the polynucleic acid moleculecomprises at least one 2′ modified nucleotide, at least one modifiedinternucleotide linkage, or at least one inverted abasic moiety, andwherein the polynucleic acid molecule is from about 10 to about 50nucleotides in length.
 2. The polynucleic acid molecule of claim 1,wherein the polynucleic acid molecule hybridizes to a target sequenceselected from SEQ ID NOs: 1-376 with less than 3 mismatched bases, lessthan 2 mismatched bases, or less than 1 mismatched bases.
 3. Thepolynucleic acid molecule of claim 1, wherein the at least one 2′modified nucleotide comprises 2′-O-methyl, 2′-O-methoxyethyl (2′-O-MOE),2′-O-aminopropyl, 2′-deoxy, T-deoxy-2′-fluoro, 2′-O-aminopropyl(2′-O-AP), 2′-O-dimethylaminoethyl (2′-O-DMAOE),2′-O-dimethylaminopropyl (2′-O-DMAP), T-O-dimethylaminoethyloxyethyl(2′-O-DMAEOE), or 2′-O—N-methylacetamido (2′-O-NMA) modified nucleotide.4. The polynucleic acid molecule of claim 1, wherein the at least one 2′modified nucleotide comprises locked nucleic acid (LNA) or ethylenenucleic acid (ENA).
 5. The polynucleic acid molecule of claim 1, whereinthe at least one inverted basic moiety is at at least one terminus. 6.The polynucleic acid molecule of claim 1, wherein the at least onemodified internucleotide linkage comprises a phosphorothioate linkage ora phosphorodithioate linkage.
 7. The polynucleic acid molecule of claim1, wherein the polynucleic acid molecule is from about 10 to about 30nucleotides in length.
 8. The polynucleic acid molecule of claim 1,wherein the polynucleic acid molecule is at least 16, 17, 18, 19, 20,21, 22, 23, 24, or 25 nucleotides in length.
 9. The polynucleic acidmolecule of claim 1, wherein the polynucleic acid molecule comprises atleast one of: from about 5% to about 100% modification, from about 10%to about 100% modification, from about 20% to about 100% modification,from about 30% to about 100% modification, from about 40% to about 100%modification, from about 50% to about 100% modification, from about 60%to about 100% modification, from about 70% to about 100% modification,from about 80% to about 100% modification, and from about 90% to about100% modification.
 10. The polynucleic acid molecule of claim 1, whereinthe polynucleic acid molecule comprises about 1, about 2, about 3, about4, about 5, about 6, about 7, about 8, about 9, about 10, about 11,about 12, about 13, about 14, about 15, about 16, about 17, about 18,about 19, about 20, about 21, about 22, or more modified nucleotides.11. The polynucleic acid molecule of claim 1, wherein the polynucleicacid molecule comprises a single strand.
 12. The polynucleic acidmolecule of claim 1, wherein the polynucleic acid molecule comprises afirst polynucleotide and a second polynucleotide hybridized to the firstpolynucleotide to form a double-stranded polynucleic acid molecule. 13.The polynucleic acid molecule of claim 12, wherein the secondpolynucleotide comprises at least one modification.
 14. The polynucleicacid molecule of claim 12, wherein the first polynucleotide and thesecond polynucleotide are RNA molecules.
 15. The polynucleic acidmolecule of claim 12, wherein the first polynucleotide comprises asequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100%sequence identity to a sequence selected from SEQ ID NOs: 377-1892. 16.The polynucleic acid molecule of claim 12, wherein the secondpolynucleotide comprises a sequence having at least 80%, 85%, 90%, 95%,96%, 97%, 98%, 99%, or 100% sequence identity to a sequence selectedfrom SEQ ID NOs: 377-1892.
 17. A pharmaceutical composition comprising:a) a molecule of claim 1; and b) a pharmaceutically acceptableexcipient.
 18. The pharmaceutical composition of claim 17, wherein thepharmaceutical composition is formulated as a nanoparticle formulation.19. The pharmaceutical composition of claim 17, wherein thepharmaceutical composition is formulated for parenteral, oral,intranasal, buccal, rectal, or transdermal administration.
 20. Apolynucleic acid molecule that mediates RNA interference against EGFR,wherein the polynucleic acid molecule comprises at least 80% sequenceidentity to a sequence selected from SEQ ID NOs: 377-1892.
 21. Thepolynucleic acid molecule of claim 20, wherein the polynucleic acidmolecule comprises at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100%sequence identity to a sequence selected from SEQ ID NOs: 377-1892. 22.The polynucleic acid molecule of claim 20, wherein the polynucleic acidmolecule is from about 10 to about 30 nucleotides in length.
 23. Thepolynucleic acid molecule of claim 20, wherein the polynucleic acidmolecule is at least 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25nucleotides in length.
 24. The polynucleic acid molecule of claim 20,wherein the polynucleic acid molecule comprises at least one of: fromabout 5% to about 100% modification, from about 10% to about 100%modification, from about 20% to about 100% modification, from about 30%to about 100% modification, from about 40% to about 100% modification,from about 50% to about 100% modification, from about 60% to about 100%modification, from about 70% to about 100% modification, from about 80%to about 100% modification, and from about 90% to about 100%modification.
 25. The polynucleic acid molecule of claim 20, wherein thepolynucleic acid molecule comprises about 1, about 2, about 3, about 4,about 5, about 6, about 7, about 8, about 9, about 10, about 11, about12, about 13, about 14, about 15, about 16, about 17, about 18, about19, about 20, about 21, about 22, or more modified nucleotides.
 26. Amethod of treating a disease or disorder in a patient in need thereof,comprising administering to the patient a composition comprising amolecule of claim
 1. 27. The method of claim 26, wherein the disease ordisorder is a cancer.
 28. The method of claim 27, wherein the cancercomprises a EGFR-associated cancer.
 29. The method of claim 27, whereinthe cancer comprises bladder cancer, breast cancer, colorectal cancer,endometrial cancer, esophageal cancer, glioblastoma multiforme, head andneck cancer, kidney cancer, lung cancer, ovarian cancer, pancreaticcancer, prostate cancer, or thyroid cancer.
 30. The method of claim 27,wherein the cancer comprises acute myeloid leukemia, CLL, DLBCL, ormultiple myeloma.
 31. A method of inhibiting the expression of EGFR genein a primary cell of a patient, comprising administering a molecule ofclaim 1 to the primary cell.
 32. The method of claim 31, wherein themethod is an in vivo method.
 33. The method of claim 31, wherein thepatient is a human.